Urinary D-glucaric acid and serum hepatic enzyme levels in chronic alcoholics

Urinary D-glucaric acid (DGA) and the activities of gamma-glutamyl transferase (GGT) and other hepatic enzymes in serum were determined in 33 noncirrhotic male alcoholics who had continued to consume alcohol until at least 24 h prior to the taking of samples. DGA excretion was significantly greater in them than in a group of 30 healthy controls (p less than 0.001), exceeding the upper reference level in 38% of the alcoholic cases (as compared with 88% for GGT). In the alcoholic patients, there was highly significant correlation between urinary DGA and serum GGT (r = 0.613, p less than 0.001), suggesting that in both cases the increased levels are due to enzyme induction. None of the biochemical variables studied were significantly correlated with estimated daily alcohol consumption. Urinary DGA levels fell off rapidly with abstinence, and in 31 alcoholic patients who had consumed no alcohol for 5 days, there was no statistically significant correlation between DGA excretion and serum GGT (r = 0.158, p congruent to 0.4).     

Expression of Intercellular Adhesion Molecule-1 (ICAM-1) on Cultured Human Endometrial Stromal Cells and Its Role in the Interaction With Natural Killers

PROBLEM: Recent evidence emphasizes the role of natural killer cells (NKs) as potential effectors of peritoneal immune surveillance directed against the outgrowth of endometrial cells, refluxed with menstrual debris, in ectopic sites. This NK-mediated cytotoxicity toward autologous endometrial antigens seems to be significantly decreased in endometriosis patients. METHOD: We set up experiments to clarify which molecules are involved in NK-endome-trial cell interaction. In particular, we evaluated the surface expression and functional activity of intercellular adhesion molecule-1 (ICAM-1), a cell surface glycoprotein that has been identified as one of the ligands for lymphocyte function-associated antigen-1 (LFA-1), present on almost all leucocyte cell types. Immunofluorescence flow cytometry was used to assess ICAM-1 expression on resting and IL 1β-activated endometrial stromal cells in culture. Dermal fibroblasts were used as control cells. Cytotoxicity and binding assays by ⁵¹Cr release in presence and absence of a specific monoclonal antibody (mAb) against ICAM-1 were then performed in order to determine the effect of this molecule on NK-mediated cytotoxic and binding activity toward endometrial stromal cells. RESULTS: The results of this study indicated that ICAM-1 expression on endometrial stromal cells seems to be constitutively higher than on dermal fibroblasts and can be up-regulated upon exposure to IL 1β. Furthermore, a mAb against ICAM-1 strongly inhibits the binding but not the cytotoxicity of NKs toward endometrial cells. No difference in the expression of this molecule was observed throughout the cycle. CONCLUSIONS: The presence of ICAM-1 on human endometrium might relate to the action of the immunocompetent cells in human specific reproductive events.     

Color-doppler velocimetry of uterine arteries in pregnant and nonpregnant patients during multiovulation induction for IVF

Objectives To evaluate uterine artery resistance during multiovulation induction in relation to the implantation rate in patients attendingin vitro fertilization (IVF) cycles.Patients Multiovulation induction for IVF was monitored by daily determination of the pulsatility index (PI) of the uterine arteries, obtained by a transvaginal probe (6.5 MHz) implemented with color-flow imaging. Doppler data were obtained from 5 days before hCG administration to the day of follicular aspiration. One IVF cycle was monitored in 70 patients. In 17 patients, 41 IVF cycles were monitored until a successful attempt occurred.Results In the 70 patients studied during one IVF attempt, the PI of the uterine arteries significantly varied (P < 0.001) in the different phases of the cycle. In the 24 patients who conceived, a significantly lower PI (P < 0.03) was found throughout the cycle. This result was mainly due to a highly significant difference of PI values observed the day after hCG administration (P < 0.005). In the 17 patients who conceived after 1 to 4 negativein vitro fertilizations, no significant difference in PI was observed in the uterine artery resistance in cycles in which implantation was or was not successful.Conclusions Uterine artery resistance varies significantly during phases of the induction therapy. Uterine artery resistance is lower throughout the course of multiovulation induction in patients with higher pregnancy rates. The PI on the day after hCG administration was the best index of pregnancy rate. Low uterine artery resistance was present even in negative attempts in patients who eventually achieved a successful implantation. PI values 3 can be considered a favorable prognostic factor for future IVF cycles.Presented at the 49th Annual Meeting of the American Fertility Society, Montreal, 1993 and the 50th Annual Meeting of the American Fertility Society, November 5–10, 1994, San Antonio, Texas.     

Predictors of failure after fecal microbiota transplantation for recurrent Clostridioides difficile infection: a systematic review and meta-analysis

Fecal microbiota transplantation (FMT) is a highly effective therapy for recurrent Clostridioides difficile infection (CDI), with ~15% 1-year recurrence rate. Small studies have identified variable risk factors associated with FMT failure. We, therefore, performed a systematic review and meta-analysis to evaluate the predictors of FMT failure. A systematic search of Medline, Embase, and Web of Science was performed from January 2013 up to June 2020. Meta-analyses were performed using random-effects models and pooled adjusted odds ratios for risk factors reported in ≥2 studies were calculated. Overall, 2671 patients with recurrent CDI who underwent FMT in 12 studies were included. FMT failure occurred in 454 patients (16.9%) with median follow-up of 3 months (range 2–7.7 months). A total of 9 risk factors were identified in ≥2 studies. Meta-analysis showed that use of non- CDI antibiotics, presence of inflammatory bowel disease, poor quality of bowel preparation, CDI-related hospitalization before FMT, inpatient FMT, and severe CDI were associated with statistically significant increased risk of failure after FMT. Increasing age, female gender, and immunocompromised status were not associated with increased risk for FMT failure. Several risk factors (both modifiable and non-modifiable) are associated with FMT failure. Lower use of antibiotics in the post-FMT period and good bowel preparation at the time of FMT are associated with lower risk of failure after FMT. Additionally, patients with non-modifiable risk factors should be counseled to be particularly alert about recurrent symptoms after FMT.

     

Plasma beta-endorphin, beta-lipotropin, and met-enkephalin concentrations during pregnancy in normal and drug-addicted women and their newborn

Most studies of plasma beta-endorphin concentrations in pregnant women show that these are highly elevated. This might indicate a role for opiate peptides during pregnancy and in the fetus-mother relationship. We measured plasma beta-endorphin, beta-lipotropin, and met-enkephalin concentrations in normal and drug-addicted women during pregnancy, labor, and delivery, and in their newborn infants. Peptides were measured by RIA after extraction and concentration on silica columns and separation by high pressure liquid chromatography. In both normal and drug-addicted mothers we found an increase in plasma beta-endorphin during pregnancy, without a concomitant increase in plasma beta-lipotropin or metenkephalin. Only beta-lipotropin increased dramatically in both groups at delivery, whereas beta-endorphin and met-enkephalin remained unchanged. Peptide concentrations in umbilical plasma were similar to those in peripheral plasma of the mothers. On day 1 of life plasma beta-endorphin, beta-lipotropin, and met-enkephalin concentrations in the newborn from normal mothers were higher than in nonpregnant adult subjects and gradually decreased toward normal adult values by day 5 of life. Plasma beta-endorphin, beta-lipotropin, and met-enkephalin concentrations of newborn infants of drug-addicted mothers increased dramatically on day 2 and 3 of life, up to 1000-fold the concentrations of normal adults, and remained elevated up to 40 days after birth. In conclusion, beta-endorphin, beta-lipotropin, and met-enkephalin concentrations during pregnancy are not affected by drug addiction, whereas in the newborn of drug addicted mothers concentrations of these compounds are markedly increased.     

Phosphorylation releases constraints to domain motion in ERK2

Protein motions control enzyme catalysis through mechanisms that are incompletely understood. Here NMR ¹³C relaxation dispersion experiments were used to monitor changes in side-chain motions that occur in response to activation by phosphorylation of the MAP kinase ERK2. NMR data for the methyl side chains on Ile, Leu, and Val residues showed changes in conformational exchange dynamics in the microsecond-to-millisecond time regime between the different activity states of ERK2. In inactive, unphosphorylated ERK2, localized conformational exchange was observed among methyl side chains, with little evidence for coupling between residues. Upon dual phosphorylation by MAP kinase kinase 1, the dynamics of assigned methyls in ERK2 were altered throughout the conserved kinase core, including many residues in the catalytic pocket. The majority of residues in active ERK2 fit to a single conformational exchange process, with k_ex ≈ 300 s⁻¹ (k_AB ≈ 240 s⁻¹/k_BA ≈ 60 s⁻¹) and p_A/p_B ≈ 20%/80%, suggesting global domain motions involving interconversion between two states. A mutant of ERK2, engineered to enhance conformational mobility at the hinge region linking the N- and C-terminal domains, also induced two-state conformational exchange throughout the kinase core, with exchange properties of k_ex ≈ 500 s⁻¹ (k_AB ≈ 15 s⁻¹/k_BA ≈ 485 s⁻¹) and p_A/p_B ≈ 97%/3%. Thus, phosphorylation and activation of ERK2 lead to a dramatic shift in conformational exchange dynamics, likely through release of constraints at the hinge.The MAP kinase, extracellular signal-regulated kinase 2 (ERK2), is a key regulator of cell signaling and a model for protein kinase activation mechanisms (1). ERK2 can be activated by MAP kinase kinases 1 and 2 (MKK1 and 2) through dual phosphorylation of Thr and Tyr residues located at the activation loop (Thr183 and Tyr185, numbered in rat ERK2) (1, 2). Phosphorylation at both sites is required for kinase activation, resulting in increased phosphoryl transfer rate and enhanced affinity for ATP and substrate (3).Conformational changes accompanying the activation of ERK2 have been documented by X-ray structures of the inactive, unphosphorylated (0P-ERK2) and the active, dual-phosphorylated (2P-ERK2) forms (4, 5). Phosphorylation rearranges the activation loop, leading to new ion-pair interactions between phospho-Thr and phospho-Tyr residues and basic residues in the N- and C-terminal domains of the kinase core structure. This leads to a repositioning of active site residues surrounding the catalytic base, enabling recognition of the Ser/Thr-Pro sequence motif at phosphorylation sites and exposing a recognition site for interactions with docking sequences in substrates and scaffolds (6).Less is known about how changes in internal motions contribute to kinase activation. Previous studies using hydrogen-exchange mass spectrometry (HX-MS) and electron paramagnetic resonance spectroscopy (7–9) led to a model where conformational mobility at the hinge linking the N- and C-terminal domains is increased by phosphorylation, therefore releasing constraints needed for activation. Such a model differs from other types of autoinhibitory mechanisms in protein kinases, which involve interactions with domains outside the kinase core (10, 11). However, how hinge flexibility regulates ERK2 is unknown.NMR relaxation dispersion methods enable protein dynamics to be monitored by measuring exchange between conformational states (12). In particular, Carr–Purcell–Meiboom–Gill (CPMG) relaxation dispersion experiments report on motions on slow (100–2,000 s⁻¹) timescales (13), which are often important for enzymatic function (13–16). In the CPMG experiment, exchange between different conformational states is probed with varying times between “refocusing” pulses. Conformational exchange leads to imperfect refocusing, thus decreasing the intensity of the NMR signal. Increasing the pulse frequency allows less chance for conformational exchange, and therefore increased NMR signal intensity. For a given pulse frequency, analysis of the signal intensity yields the effective relaxation rate for the resonance, R_2,eff. This is typically plotted as a relaxation dispersion curve, which can be fit to a two-state conformational exchange process (e.g., A ⇌ B interconversion). Fitting extracts the populations and the exchange rates between states, thus reflecting the thermodynamics and kinetics of the system (17, 18).Here we performed CPMG relaxation dispersion experiments at multiple field strengths to compare the dynamic properties of [¹³C]methyl-labeled ERK2 in its phosphorylated and unphosphorylated states. The results demonstrate that phosphorylation causes a significant change in exchange dynamics throughout the kinase core, consistent with a global domain motion. Increasing hinge mobility by introducing mutations at the hinge also promotes domain motion within the core but with differing kinetics and populations. Taken together, the results show that large changes in dynamics accompany ERK2 phosphorylation, which are influenced by conformational mobility at the hinge. We propose that the activation of ERK2 involves removing inhibitory constraints to domain motion, which are conferred by the internal architecture of the kinase.     

Gastro 2013 APDW/WCOG Shanghai Working Party Report: Chronic diarrhea: Definition,classification, diagnosis

Diarrhea is best defined as passage of loose stools often with more frequent bowel movements. For clinical purposes, the Bristol Stool Form Scale works well to distinguish stool form and to identify loose stools. Laboratory testing of stool consistency has lagged behind. Acute diarrhea is likely to be due to infection and to be self‐limited. As diarrhea becomes chronic, it is less likely to be due to infection; duration of 1 month seems to work well as a cut‐off for chronic diarrhea, but detailed scientific knowledge is missing about the utility of this definition. In addition to duration of diarrhea, classifications by presenting scenario, by pathophysiology, and by stool characteristics (e.g. watery, fatty, or inflammatory) may help the canny clinician refine the differential diagnosis of chronic diarrhea. In this regard, a careful history remains the essential part of the evaluation of a patient with diarrhea. Imaging the intestine with endoscopy and radiographic techniques is useful, and biopsy of the small intestine and colon for histological assessment provides key diagnostic information. Endomicroscopy and molecular pathology are only now being explored for the diagnosis of chronic diarrhea. Interest in the microbiome of the gut is increasing; aside from a handful of well‐described infections because of pathogens, little is known about alterations in the microbiome in chronic diarrhea. Serological tests have well‐defined roles in the diagnosis of celiac disease but have less clearly defined application in autoimmune enteropathies and inflammatory bowel disease. Measurement of peptide hormones is of value in the diagnosis and management of endocrine tumors causing diarrhea, but these are so rare that these tests are of little value in screening because there will be many more false‐positives than true‐positive results. Chemical analysis of stools is of use in classifying chronic diarrhea and may limit the differential diagnosis that must be considered, but interpretation of the results is still evolving. Breath tests for assessment of carbohydrate malabsorption, small bowel bacterial overgrowth, and intestinal transit are fraught with technical limitations that decrease sensitivity and specificity. Likewise, tests of bile acid malabsorption have had limited utility beyond empirical trials of bile acid sequestrants.     

Quantitative variation in lysosomes in the epithelium of the human uterine tube in the menstrual cycle and after the menopause]

Quantitative variation of the lysosomes in the epithelium of the human uterine tube in the menstrual cycle and in post-menopausal period. The authors counted the lysosomes in the human uterine tube in the menstrual cycle and in post-menopausal period. The post-menopausal women were separated in two groups: one received oral estrogens during twenty one day and the other did not. Fragments of uterine ampullae were removed, fixed and submitted to Gomori acid phosphatase method. Results show that the number of lysosomes during the proliferative phase was significantly greater than in secretory phase and post-menopausal period. The number of lysosomes in post-menopausal period was significantly greater in women treated with estrogens. Our result show that estrogens are responsible for the increase of the number of lysosomes.