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This study compares wrist motion, biomechanical behaviour and radiographic parameters before and after total wrist arthroplasty using a fourth-generation spherical articulation prosthesis. A total of 10 cadaveric specimens were assessed using a hexapod Stewart platform robot. After arthroplasty, there were significant increases in both stiffness and phase angle of wrist motion across all planes of motion assessed. In three specimens, a sudden increase in moment was observed on load/displacement curves. Radiographically, carpal height increased by 14%, and the centre of rotation was displaced 11.1 mm proximally, 4.6 mm dorsally and 3.9 mm radially. This stretched the musculotendinous units, tightening the joint, while increasing the moment arm of the wrist flexors and decreasing the moment arm of the extensors, potentially important in the development of postoperative flexion contractures. Possible alterations in technique and/or implant design are considered to assist surgeons in achieving optimal clinical and survivorship outcomes., Competing Interests: Declaration of conflicting interestsThe authors declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Background: Facet fractures are frequently associated with clinically observed cervical facet dislocations (CFDs); however, to date there has only been one experimental study, using functional spinal units (FSUs), which has systematically produced CFD with concomitant facet fracture. The role of axial compression and distraction on the mechanical response of the cervical facets under intervertebral motions associated with CFD in FSUs has previously been shown. The same has not been demonstrated in multi-segment lower cervical spine specimens under flexion loading (postulated to be the local injury vector associated with CFD)., Methods: This study investigated the mechanical response of the bilateral inferior C6 facets of thirteen C5-C7 specimens (67±13 yr, 6 male) during non-destructive constrained flexion, superimposed with each of five axial conditions: (1) 50 N compression (simulating weight of the head); (2-4) 300, 500, and 1000 N compression (simulating the spectrum of intervertebral compression resulting from neck muscle bracing prior to head-first impact and/or externally applied compressive forces); and, (5) 2 mm of C6/C7 distraction (simulating the intervertebral distraction present during inertial loading of the cervical spine by the weight of the head). Linear mixed-effects models (α = 0.05) assessed the effect of axial condition., Results: Increasing amounts of intervertebral compression superimposed on flexion rotations, resulted in increased facet surface strains (range of estimated mean difference relative to Neutral: maximum principal = 77 to 110 με, minimum principal = 126 to 293 με, maximum shear = 203 to 375 με) and angular deflection of the bilateral inferior C6 facets relative to the C6 vertebral body (range of estimated mean difference relative to Neutral = 0.59° to 1.47°)., Conclusions: These findings suggest increased facet engagement and higher load transfer through the facet joint, and potentially a higher likelihood of facet fracture under the compressed axial conditions., Competing Interests: The authors declare that no benefits in any form have been or will be received from a commercial party related directly or indirectly to the subject of this manuscript., (© 2024 The Author(s). JOR Spine published by Wiley Periodicals LLC on behalf of Orthopaedic Research Society.)

Purpose of Review: This review aims to explore in-depth the different aspects of the association between very low-calorie ketogenic diet (VLCKD), obesity and obesity-related thyroid dysfunction., Recent Findings: The VLCKD, proposed as a non-pharmacological strategy for the management of certain chronic diseases, is becoming increasingly popular worldwide. Initially used to treat epilepsy, it has been shown to be effective in controlling body weight gain and addressing various pathophysiological conditions. Research has shown that a low-calorie, high-fat diet can affect thyroid hormone levels. Weight loss can also influence thyroid hormone levels. Studies have suggested that long-term use of VLCKD for refractory epilepsy may be related to the development of hypothyroidism, with an effect seen in various populations. In particular, women with obesity following VLCKD tend to have reduced T3 levels. We propose further research to unravel the underlying mechanisms linking VLCKD to obesity and obesity-related thyroid dysfunction., (© 2024. The Author(s).)

Objective: To estimate the proportion of individuals with established Type 2 Diabetes Mellitus (T2DM) and Cardiovascular Disease (CVD) who are receiving pharmacological anti-diabetic treatment with evidence of cardiovascular benefit at a hospital in Argentina., Materials and Methods: Cross-sectional study conducted at the Italian Hospital of Buenos Aires. A consecutive sample of adult patients affiliated with the institutional prepaid health plan active in March 2020, diagnosed with T2DM and established CVD, was included. Data were collected from the Electronic Health Record. The proportion of pharmacological adequacy (combined use of metformin plus sodium-glucose co-transporter 2 inhibitors and/or glucagon-like peptide 1 receptor agonists) was reported along with its respective 95% confidence interval (CI)., Results: A total of 1539 patients were included, with a mean age of 76.2 years; 65.3% were male, and 81.6% were overweight or obese. Hemoglobin A1c levels were recorded in the past year for 74.9% of patients, with an average value of 6.9% (SD 1.2). The most prescribed drugs were metformin (61.3%), insulin (26.7%), and gliptins (11%). Out of the total included patients, 82 exhibited pharmacotherapeutic adequacy for diabetes treatment, with a prevalence of 5.3% (95% CI 4.2-6.5)., Conclusions: The prevalence of prescribing anti-diabetic drugs with evidence of cardiovascular benefit was 5.3% (95% CI 4.2-6.5). This real-world evidence highlights the low frequency of prescribing this type of medication at the time of the study in a high cardiovascular risk population., (Universidad Nacional de Córdoba)

Background: Hospital hypoglycemia (HH) is an important problem since it is linked to hospital death, prolonged stay and higher costs, both in people with and without diabetes, in the critical area and general ward, especially in cases of severe hypoglycemia. The objective of our work was to compare hospital mortality, transfer to the critical area and hospitalization periods in adults with HH according to its severity., Methods: We conducted a retrospective cohort of adults hospitalized with hypoglycemia at the Hospital Italiano de Buenos Aires between 2013 and 2018. Three groups of hypoglycemia were defined: mild (glycemia between 70 and 54 mg/dl), severe (glycemia < 54 mg/dl and = 40 mg/dl) and critical (glycemia < 40mg/dl)., Results: A total of 5994 patients had HH, the majority presented mild hypoglycemia (72%). Severe and critical hypoglycemia, compared with mild hypoglycemia, were associated with higher mortality: OR 2.6 (95%CI 2.2-3.1) and 4.2 (95%CI 3.5-5.0) respectively; and increased hospitalization in critical care areas: OR 1.6 (95%CI 1.4-1.9) and 3.2 (95%CI 2.6-4.0) respectively. There were no differences in length of stay., Discussion: This information provides, in our country, evidence on the prognostic value of hypoglycemia, whose greater severity is associated with increased hospitalization in critical areas and higher hospital mortality.

Objective: To estimate prevalence of diabetes in outpatient care and to describe its epidemiological characteristics, comorbidities, and related vascular complications., Methods: Observational cross-sectional study which included all adults affiliated from a private insurance health plan on March 2019, at Hospital Italiano de Buenos Aires, from Argentina., Results: The global prevalence of diabetes resulted in 8.5% with 95% CI 8.3-8.6 (12,832 out of a total of 150,725 affiliates). The age stratum with the highest prevalence was the group between 65 and 80 years old with 15.7% (95% CI 15.3-16.1). People with diabetes had a mean age of 70 years (SD 14), 52% were women, and the most frequently associated cardiovascular risk factors were: dyslipidaemia (88%), arterial hypertension (74%) and obesity (55%). In relation to metabolic control, 60% had at least one glycosylated hemoglobin measured in the last year, 70% of which were less than 7%. Almost 80% have LDL measured at least once in the last 2 years, 55% of them had an LDL value equal to or less than 100 mg/dl. The macrovascular complications present in order of frequency were: acute myocardial infarction (11%), cerebrovascular accident (8%) and peripheral vascular disease (4%); while the microvascular complications were found to be diabetic neuropathy (4%) and retinopathy (2%). 7% had diabetic foot, with less than 1% amputations., Conclusion: Diabetes represents a prevalent problem, even in elderly patients. This population continues to present a high cardiovascular risk, with little compliance with therapeutic goals., (Copyright: © 2023 Permanyer.)

Electronic Health Records offer an opportunity to improve patient care (in terms of quality and/or safety) by making available patient health information stored in a single Clinical Data Repository. We aimed to estimate the frequency of hypoglycemic recurrences in hospitalized adult patients in non-critical areas. We designed a cross sectional study with hospitalizations between 2017 and 2018, which included patients with at least one hypoglycemia health record (defined as a value <70 mg/dL, either by capillary glycemic monitoring or serum measurement). Recurrence was defined as those who presented a second event, with at least 2 hours of difference. We included 1884 patients, and 573 presented recurrences, yielding a global prevalence of 30.41% (95%CI 28.34-32.54). Due to the high frequency, it is important to identify vulnerable populations, to implement preventive measures to assist clinicians for decision-making tasks, as a clinical decision support system.

The main objective was to estimate the prevalence of diabetes mellitus (DM) among patients with COVID-19, to explore associated factors; and to describe clinical evolution of hospitalized patients. A cross-sectional study was conducted, which included adults confirmed with COVID-19 between 03/12/2020 and 10/15/2020, at Hospital Italiano de Buenos Aires. From 6009 people with COVID-19, 408 had previous diagnosis of DM, yielding a prevalence of 6% (95%CI 6-7), higher prevalence was associated with age (12% in = 60 years and 3% in < 60 years; p = 0.01). In-hospital mortality was 6% (95%CI 6-7), being 15% in DM and 6% compared in non-diabetics (p < 0.01). Associated factors with DM were cardiovascular variables such as male sex, hypertension, smoking, chronic renal failure, heart failure, previous coronary disease; and clinical variables proxy of frailty such as: age, dementia and previous institutionalization (all with p < 0.01). Only 23% (96/408) of DM had an HbA1c measurement in the last 3 months and 76% in the last year, with an average 8.6%, and 25% in goal (HbA1c = 7%). Management was mostly in-hospital (59%), with an average hospital stay of 12 days, with the following complications during hospitalization: 6% presented a hypoglycemic value (< 70 mg/dl), 42% required oxygen therapy, 19 % went to intensive care unit, 15% required invasive mechanical ventilation (mean 11 days), and 25% (95%CI 20-31) of in-hospital mortality (mean 82 years).

The number of older adults unable to transfer or ambulate independently is increasing. High support chairs enable people experiencing loss of mobility to be mobile, but current chair designs are associated with global functional loss and pressure ulcers. This pilot study compared the functionality of a traditional design high support chair to a new design of motorised high support chair: 1) a motion laboratory study compared joint angles and pressure at the hip, knee, ankle, elbow and spine when pushing each chair, and 2) a pressure mapping study compared the interface pressure when older people with limited mobility used the chairs. Significant reduction in joint angles for the person pushing the chair (degree difference range -3.6 to 14.2) and decreased seated pressure (w/kg difference range -0.2 to 2.1) for the seated user were identified for the motorised chair. Longitudinal investigations are required to determine if the significant differences identified in this pilot study result in less manual handling injuries and pressure areas., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

BACKGROUND: Stress-induced hyperglycemia is a phenomenon that occurs typically in patients hospitalized for acute disease and resolves spontaneously after regression of the acute illness. However, it can also occur in diabetes patients, a fact that is sometimes overlooked. It is thus important to make a proper diabetes diagnosis if hospitalized patients with episodes of hyperglycemia with and without diabetes are studied. AIMS: To estimate the extent of the association between stress-induced hyperglycemia and in-hospital mortality in patients with hospital hyperglycemia (HH), and to explore potential differences between patients diagnosed with diabetes (HH-DBT) and those with stress-induced hyperglycemia (SH), but not diagnosed with diabetes. METHODS: A cohort of adults with hospital hyperglycemia admitted to a tertiary, university hospital in Buenos Aires, Argentina, was analyzed retrospectively. RESULTS: In the study, 2,955 patients were included and classified for analysis as 1,579 SH and 1,376 HH-DBT. Significant differences were observed in glycemic goal (35.53% SH versus 25.80% HH-DBT, p < 0.01), insulin use rate (26.66% SH versus 46.58% HH-DBT, p < 0.01), and severe hypoglycemia rate (1.32% SH versus 1.74% HH-DBT, p < 0.01). There were no differences in hypoglycemia rate (8.23% SH versus 10.53% HH-DBT) and hospital mortality. There was no increase in risk of mortality in the SH group adjusted for age, non-scheduled hospitalization, major surgical intervention, critical care, hypoglycemia, oncological disease, cardiovascular comorbidity, and prolonged hospitalization. CONCLUSIONS: In this study, we observed better glycemic control in patients with SH than in those with HH-DBT, and there was no difference in hospital mortality.

Sweet oranges are an important source of ascorbic acid (AsA). In this study, the content of AsA in the juice and leaves of four orange clonal selections, different in terms of maturity time and the presence/absence of anthocyanins, was correlated with the transcription levels of the main genes involved in the biosynthesis, recycling, and degradation pathways. Within each variety, differences in the above pathways and the AsA amount were found between the analysed tissues. Variations were also observed at different stages of fruit development and maturation. At the beginning of fruit development, AsA accumulation was attributable to the synergic action of l-galactose and Myo-inositol, while the l-gulose pathway was predominant between the end of fruit development and the beginning of ripening. In leaves, the l-galactose pathway appeared to play a major role in AsA accumulation, even though higher GalUr isoform expression suggests a synergistic contribution of both pathways in this tissue. In juice, the trend of the AsA content may be related to the decrease in the transcription levels of the GME , GDH , MyoOx , and GalUr12 genes. Newhall was the genotype that accumulated the most AsA. The difference between Newhall and the other varieties seems to be attributable to the GLDH , GalUr12 , APX2 , and DHAR3 genes.

Background: Anthocyanin pigmentation characterizes a number of tissues of Citrus and its relatives. The gain and loss of pigmentation is intriguing and is inherited variously among species., Methods: Citrus germplasm was used to investigate the anthocyanin pigmentation of tissues never before considered, including stamen, style and stigma, and of young leaves, petals, rind and flesh of 28 genotypes belonging to 14 species. Citrus genotypes encompassed citron, lemon, sweet orange, lime, and Citrus relatives included Microcitrus , Murraya , and Severinia . A relative qRT-PCR analysis was carried out on the structural and regulatory genes: phenylalanine ammonia-lyase (PAL), chalcone synthase (CHS), chalcone isomerase (CHI), flavanone 3'-hydroxylase (F3H), dihydroflavonol 4-reductase (DFR), anthocyanidin synthase (ANS), uridine diphosphate glucose flavonoid glucosyl-transferase (UFGT), glutathione S-transferase (GST), Ruby and Noemi. Image analysis and a genomic approach were employed to evaluate how the red pigmentation is inherited among tissues and species., Results: Pigmentation of young leaves and petals is specific to citron and its hybrids. Ruby controls the pigmentation of petals, but not of leaves. The red color of the rind and flesh is a trait that particularly characterizes a diversity of sweet oranges, citron hybrids and Citrus relatives. Color expression depends on external factors and also on developmental stage. The coloration of stamen and style is citron-specific, while a red stigma is exclusive to Moro orange and its hybrids., Conclusion: It is hypothesized that there is a relationship among Citrus species and genes controlling anthocyanin pigmentation.

Introduction: The study objective was to estimate during post-discharge follow-up the incidence of diabetes and to ascertain mortality in hospitalized patients, classified during follow-up as having stress hyperglycemia (SH) or normoglycemia (NG) based on blood glucose levels., Material and Methods: A retrospective cohort of non-diabetic adults with SH (> 140mg/dl and HbA1c <6.5%) or NG (all blood glucose values ≤ 140mg/dl) was used., Results: There were 3981 patients with NG and 884 with SH. During the observation period (median follow-up of 1.83 years), there were 255 cases of diabetes and 831 deaths. The cumulative incidence of diabetes per year was 1.59% (95% CI: 1.23-2.06) in patients with NG and 7.39% (95% CI: 5.70-9.56) in those with SH. SH was significantly associated to diabetes (crude HR 1.33, 95% CI: 1.13-1.73, p .025), even after adjusting for age and sex (adjusted HR 1.38, 95% CI 1.06-1.78, p .014). The mortality rate at one year was 10.07% (95% CI: 9.18-11.05) in NG patients and 13.24% (95% CI: 11.17-15.65) in SH patients. The sub-hazard ratio of developing diabetes considering death as a competitive event was 1.41 (95% CI 1.29-1.53, p <.001)., Conclusions: SH is a risk factor for diabetes. There were no differences in mortality during follow-up, but death appears to be a competitive event in development of diabetes in this population., (Copyright © 2018 SEEN y SED. Publicado por Elsevier España, S.L.U. All rights reserved.)

Citrus tristeza virus (CTV) is the causal agent of various diseases with dramatic effects on citrus crops worldwide. Most Citrus species, grown on their own roots, are symptomless hosts for many CTV isolates. However, depending on different scion-rootstock combination, CTV infection should result in distinct syndromes, being 'tristeza' the more severe one, leading to a complete decline of the susceptible plants in a few weeks. Transcriptomic analyses revealed several genes involved either in defense response, or systemic acquired resistance, as well as transcription factors and components of the phosphorylation cascades, to be differentially regulated during CTV infection in Citrus aurantifolia species. To date little is known about the molecular mechanism of this host-pathogen interaction, and about the rootstock effect on citrus response to CTV infection. In this work, the response to CTV infection has been investigated in tolerant and susceptible scion-rootstock combinations by two-dimensional gel electrophoresis (2DE). A total of 125 protein spots have been found to be differently accumulated and/or phosphorylated between the two rootstock combinations. Downregulation in tolerant plants upon CTV infection was detected for proteins involved in reactive oxygen species (ROS) scavenging and defense response, suggesting a probable acclimation response able to minimize the systemic effects of virus infection. Some of these proteins resulted to be modulated also in absence of virus infection, revealing a rootstock effect on scion proteome modulation. Moreover, the phospho-modulation of proteins involved in ROS scavenging and defense response, further supports their involvement either in scion-rootstock crosstalk or in the establishment of tolerance/susceptibility to CTV infection., (© 2015 Scandinavian Plant Physiology Society.)

Two cases of granulocytic sarcoma (GS) with lymph node presentation without marrow involvement are presented because it is a rare presentation of acute myeloid leukemia (AML) that can coexist with or precede it and as initial symptom (case 1) or disease relapse (case 2). The most common differential diagnoses are lymphoma or solid tumor. Biopsy is essential for definitive diagnosis. Imaging studies such as positron emission tomography (PET) are very useful for staging and monitoring. The prognosis and treatment according to the literature is not different from AML. The two cases presented were diagnosed with lymph node biopsy with a bone marrow that did not meet criteria for AML.

Four cases of a rare paraneoplastic syndrome associated to a plasmatic cell disorder with polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy and skin lesions (POEMS) are here reported. The purpose of the communication is to warn of different forms of presentation of POEMS syndrome to decrease the time of diagnosis, because early treatment reduces sequels and improves quality of life in the long term, also to remark the importance of classifying the hematological disease for specific treatment.

A flesh-specific oligonucleotide custom array was designed to study gene expression during blood orange ripening. The array included 301 probes derived from a subtracted SSH library, a cDNA-AFLP collection, and a set of regulatory genes from the Harvest citrus database. The custom array was hybridized using samples of Moro, a pigmented cultivar, and Cadenera, a common cultivar, at three different ripening stages: the immature phase, the halfway point of maturation (corresponding to the start of Moro pigmentation) and the full ripening. Of the 301 probes, 27 in total, corresponding to 20 different transcripts, indicated differential expression in stage-to-stage and/or cultivar-to-cultivar comparisons. Transcripts encoding for anthocyanin biosynthesis represented most of the total over-expressed probes. The remaining differentially expressed transcripts were functionally associated with primary metabolism as flavor biosynthesis, defense and signal transduction. The expressed products associated with probes indicating differential expression were confirmed by qRT-PCR. The microarray was designed considering a small collection of sequences useful for monitoring specific pathways and regulatory genes related to fruit ripening and anthocyanin pigmentation. The main novelty of this customization is the use of expressed sequences specifically derived from blood orange flesh to study different cultivars and ripening stages, and the provision of further information about processes related to anthocyanin pigmentation in citrus fruit flesh., (Copyright 2009 Elsevier GmbH. All rights reserved.)

A combination of 2-DE and LC-MSMS approaches was used to identify the differentially expressed proteome of a pigmented sweet orange (Citrus sinensis, cv. Moro) in comparison with a common cultivar (Cadenera) at ripening time. The comparison of the protein patterns of Moro and Cadenera showed 64 differential expressed protein spots. Fifty-five differentially expressed proteins were identified. Proteins were classified according to their putative function and known biosynthetic pathways. Most of the proteins related to sugar metabolism were overexpressed in Moro, while those related to stress responses were overexpressed in Cadenera. The abundance of proteins belonging to Unknown/Unnamed and Hypothetical classes could be associated to the incomplete data available on the Citrus genome. The relative abundance of Secondary metabolism and Oxidative process proteins substantiated the key role of the anthocyanin pathway in Moro, which is characterized by a strong pigmentation at ripening time. The potential role of protein differential expression between Moro and Cadenera fruits was discussed, and proteomic results were compared with the known variations of transcripts of the same fruits. The latter analyses highlighted many discrepancies, confirming the necessity to associate both proteomic and transcriptomic approaches in order to achieve a more complete characterization of the biological system.

Cytokine-stimulated IkappaBalpha degradation is impaired in HT-29 and primary intestinal epithelial cells. To gain more insight into the mechanism of this defect, we dissected cytokine-induced NF-kappaB signaling pathway in HT-29 cells. IL-1beta and TNF, alone or in combination with IFNgamma, failed to induce IkappaBalpha or IkappaBbeta degradation in HT-29 cells. Despite similar 125I-IL-1beta binding, HT-29 cells displayed no IRAK degradation, a 75% reduction of IKK activity, and decreased IkappaBalpha phosphorylation, NF-kappaB DNA binding activity and IL-8 mRNA accumulation in response to IL-1beta compared to Caco-2 cells. Selective activation of NF-kappaB pathway by adenoviral delivery of NF-kappaB-inducing kinase (Ad5NIK) or IKKbeta (Ad5IKKbeta) strongly activated IKK activity (>20 fold) in HT-29 cells with concomitant endogenous IkappaBalpha serine 32 phosphorylation and total IkappaBalpha degradation. In addition, NF-kappaB DNA binding activity and IL-8 secretion is higher in Ad5NIK-infected than in IL-1beta-stimulated HT-29 cells. These data show that altered NF-kappaB signaling is associated with impaired stimulation of an upstream IKK activator.

Bacterial lipopolysaccharide (LPS) stimulates Kupffer cells and participates in the pathogenesis of alcohol-induced liver injury. However, it is unknown whether LPS directly affects hepatic stellate cells (HSCs), the main fibrogenic cell type in the injured liver. This study characterizes LPS-induced signal transduction and proinflammatory gene expression in activated human HSCs. Culture-activated HSCs and HSCs isolated from patients with hepatitis C virus-induced cirrhosis express LPS-associated signaling molecules, including CD14, toll-like receptor (TLR) 4, and MD2. Stimulation of culture-activated HSCs with LPS results in a rapid and marked activation of NF-kappaB, as assessed by in vitro kinase assays for IkappaB kinase (IKK), IkappaBalpha steady-state levels, p65 nuclear translocation, NF-kappaB-dependent luciferase reporter gene assays, and electrophoretic mobility shift assays. Lipid A induces NF-kappaB activation in a similar manner. Both LPS- and lipid A-induced NF-kappaB activation is blocked by preincubation with either anti-TLR4 blocking antibody (HTA125) or Polymyxin B. Lipid A induces NF-kappaB activation in HSCs from TLR4-sufficient (C3H/OuJ) mice but not from TLR4-deficient (C3H/HeJ) mice. LPS also activates c-Jun N-terminal kinase (JNK), as assessed by in vitro kinase assays. LPS up-regulates IL-8 and MCP-1 gene expression and secretion. LPS-induced IL-8 secretion is completely inhibited by the IkappaB super repressor (Ad5IkappaB) and partially inhibited by a specific JNK inhibitor, SP600125. LPS also up-regulates cell surface expression of ICAM-1 and VCAM-1. In conclusion, human activated HSCs utilize components of TLR4 signal transduction cascade to stimulate NF-kappaB and JNK and up-regulate chemokines and adhesion molecules. Thus, HSCs are a potential mediator of LPS-induced liver injury.

Pathogenic and enteroinvasive bacteria have been shown to trigger the I kappa B/NF-kappa B transcriptional system and proinflammatory gene expression in epithelial cells. In this study, we investigated the molecular mechanism of the commensal Gram-negative Bacteroides vulgatus-induced NF-kappa B signal transduction in intestinal epithelial cells (IEC). We report that B. vulgatus induced interleukin-1 receptor-associated kinase-1 degradation, I kappa B alpha phosphorylation/degradation, RelA and Akt phosphorylation, as well as NF-kappa B DNA binding and NF-kappa B transcriptional activity in rat non-transformed IEC-6 cells. B. vulgatus- but not interleukin-1 beta-mediated NF-kappa B transcriptional activity was inhibited by dominant negative (dn) toll-like receptor 4. Of importance, B. vulgatus induced I kappa B alpha phosphorylation/degradation and IKK alpha/beta and RelA phosphorylation in primary IEC derived from germ-free or mono-associated HLA-B27 transgenic and wild type rats, demonstrating the physiological relevance of non-pathogenic bacterial signaling in IEC. Adenoviral delivery of dn IKK beta or treatment with wortmannin inhibited B. vulgatus-induced endogenous RelA Ser-536 and GST-p65TAD (Ser-529/Ser-536) phosphorylation as well as NF-kappa B transcriptional activity in IEC-6 cells, suggesting a critical role of IKK beta and phosphatidylinositol 3-kinase/Akt in bacteria-induced RelA phosphorylation and NF-kappa B activation. Interestingly, B. vulgatus-induced I kappa B alpha degradation and NF-kappa B transcriptional activity in IEC transwell cultures were inhibited in the presence of lymphocytes. We propose that non-pathogenic B. vulgatus activates the NF-kappa B signaling pathway through both I kappa B degradation and RelA phosphorylation but that immune cells mediate tolerance of IEC to this commensal bacteria.

The function of nuclear factor (NF)-kappaB within the developing and mature CNS is controversial. We have generated transgenic mice to reveal NF-kappaB transcriptional activity in vivo. As expected, constitutive NF-kappaB activity was observed within immune organs, and tumor necrosis factor-inducible NF-kappaB activity was present in mesenchymal cells. Intriguingly, NF-kappaB activity was also prominent in the CNS throughout development, especially within neocortex, olfactory bulbs, amygdala, and hippocampus. NF-kappaB in the CNS was restricted to neurons and blocked by overexpression of dominant-negative NF-kappaB-inducible kinase or the IkappaBalphaM super repressor. Blocking endogenous neuronal NF-kappaB activity in cortical neurons using recombinant adenovirus induced neuronal death, whereas induction of NF-kappaB activity increased levels of anti-apoptotic proteins and was strongly neuroprotective. Together, these data demonstrate a physiological role for NF-kappaB in maintaining survival of central neurons.

In this study, we examined the role of the nuclear factor-kappaB (NF-kappaB)-inducing kinase (NIK) in distinct signaling pathways leading to NF-kappaB activation. We show that a dominant-negative form of NIK (dnNIK) delivered by adenoviral (Ad5dnNIK) vector inhibits Fas-induced IkappaBalpha phosphorylation and NF-kappaB-dependent gene expression in HT-29 and HeLa cells. Interleukin (IL)-1beta- and tumor necrosis factor-alpha (TNF-alpha)-induced NF-kappaB activation and kappaB-dependent gene expression are inhibited in HeLa cells but not in Ad5dnNIK-infected HT-29 cells. Moreover, Ad5dnNIK failed to sensitize HT-29 cells to TNF-alpha-induced apoptosis at an early time point. However, cytokine- and Fas-induced signals to NF-kappaB are finally integrated by the IkappaB kinase (IKK) complex, since IkappaBalpha phosphorylation, NF-kappaB DNA binding activity, and IL-8 gene expression were strongly inhibited in HT-29 and HeLa cells overexpressing dominant-negative IKKbeta (Ad5dnIKKbeta). Our findings support the concept that cytokine signaling to NF-kappaB is redundant at the level of NIK. In addition, this study demonstrates for the first time the critical role of NIK and IKKbeta in Fas-induced NF-kappaB signaling cascade.

Background and Aims: Cyclooxygenase (COX)-2 is up-regulated in most colonic cancers and in inflammatory bowel disease in which tumor necrosis factor (TNF)-alpha is believed to play a central role. There has been recent speculation on the activation of phosphatidylinositol 3-kinase (PI 3-kinase) by TNF-alpha and its role in the regulation of genes controlled by NF-kappaB. We investigated the regulatory role of PI 3-kinase on COX-2 expression in colonic epithelial cells., Methods: In HT-29 and Caco-2 colonic epithelial cells, COX-2 expression was induced by either TNF-alpha or interleukin (IL)-1alpha as observed by Northern and Western analyses. COX-2 activity was assessed by measuring prostaglandin E(2) (PGE2) production by enzyme-linked immunosorbent assay. NF-kappaB binding activity was assessed by electrophoretic mobility shift assay. PI 3-kinase activity was measured by quantifying the accumulation of PI 3-kinase-dependent D-3 lipid products by high-performance liquid chromatography., Results: The PI 3-kinase inhibitor wortmannin up-regulated induced COX-2 expression in a concentration-dependent manner in both HT-29 and Caco-2 cells. An alternative PI 3-kinase inhibitor, LY294002, caused up-regulation of induced COX-2 messenger RNA (mRNA) in HT-29 cells at concentrations of < or =1 micromol/L. IL-4 and IL-13, which are known to activate PI 3-kinase, down-regulated HT-29 COX-2 mRNA, protein, and PGE2 production. NF-kappaB binding activity was unaltered by PI 3-kinase inhibition in HT-29 cells, in which TNF-alpha was shown to activate PI 3-kinase directly., Conclusions: COX-2 is negatively regulated by PI 3-kinase; we propose that the inhibitory effect of IL-4 and IL-13 is mediated via a PI 3-kinase-dependent pathway. This mechanism does not appear to involve NF-kappaB because PI 3-kinase inhibition did not alter NF-kappaB binding activity. TNF-alpha can activate PI 3-kinase directly in addition to inducing COX-2.

NF-kappa B plays a critical role in the transcriptional regulation of proinflammatory gene expression in various cells. Cytokine-mediated activation of NF-kappa B requires activation of various kinases, which ultimately leads to the phosphorylation and degradation of I kappa B, the NF-kappa B cytoplasmic inhibitor. The food derivative curcumin has been shown to inhibit NF-kappa B activity in some cell types. In this report we investigate the mechanism of action of curcumin on cytokine-induced proinflammatory gene expression using intestinal epithelial cells (IEC). Curcumin inhibited IL-1 beta-mediated ICAM-1 and IL-8 gene expression in IEC-6, HT-29, and Caco-2 cells. Cytokine-induced NF-kappa B DNA binding activity, RelA nuclear translocation, I kappa B alpha degradation, I kappa B serine 32 phosphorylation, and I kappa B kinase (IKK) activity were blocked by curcumin treatment. Wound-induced p38 phosphorylation was not inhibited by curcumin treatment. In addition, mitogen-activated protein kinase/ERK kinase kinase-1-induced IL-8 gene expression and 12-O-tetraphorbol 12-myristate 13-acetate-responsive element-driven luciferase expression were inhibited by curcumin. However, I kappa B alpha degradation induced by ectopically expressed NF-kappa B-inducing kinase or IKK was not inhibited by curcumin treatment. Therefore, curcumin blocks a signal upstream of NF-kappa B-inducing kinase and IKK. We conclude that curcumin potently inhibits cytokine-mediated NF-kappa B activation by blocking a signal leading to IKK activity.

Interleukin-2 (IL-2) and IL-15 exert similar biological actions, which largely reflect the fact that their receptors share common beta and gamma subunits; in contrast, distinct subunits are required for high-affinity binding of either cytokine to a heterotrimeric receptor complex. Human neutrophils are known to express both the beta and gamma subunits of the IL-2/IL-15 receptor complex, and we now report that they also constitutively express messenger RNA transcripts encoding the IL-15 receptor chain, suggesting that they possess functional, heterotrimeric IL-15 receptors. Accordingly, we show that in neutrophils, IL-15 elicits several functional responses. In particular, neutrophils synthesize and release IL-8 in response to IL-15, but not to IL-2. Moreover, a nuclear factor-kappaB (NF-kappaB) DNA-binding activity was enhanced in nuclear extracts of IL-15-treated neutrophils, which could be supershifted by antibodies to p50 or RelA. Again, no detectable effect of IL-2 was observed on this response. In peripheral blood lymphocytes (PBL), however, both IL-2 and IL-15 were potent inducers of NF-kappaB activation. Conversely, neither IL-15 nor IL-2 elicited the formation of activator protein-1 (AP-1) DNA-binding complexes in neutrophils, even though both cytokines were found to activate these DNA-binding activities in PBL. Collectively, these observations establish neutrophils as a useful cellular model to discriminate between the actions of IL-15 and IL-2. More importantly, this is the first demonstration that IL-15 has the ability to induce NF-kappaB and AP-1 activation, which further emphasizes the potential relevance of this newly discovered cytokine to immune and inflammatory processes.

In neutrophils of a chronic granulomatous disease (CGD) patient with a lack of p67phox the mRNA for p67phox was present in normal amount and size. This mRNA was reverse transcribed, and the coding region was analyzed by single-strand conformation polymorphism analysis. Direct DNA sequencing allowed the identification of a A479-to-T and A481-to-G substitution in exon 5 of the p67phox gene resulting in a double nonconservative amino acid change 160Lys-to-Glu and 161Asp-to-Val (D160V-K161E). This defect was found in the genomic DNA of this patient in heterozygous state and does not correspond to those previously found in other cases of CGD lacking the p67phox.

GRO alpha, a member of the chemokine superfamily, exerts potent stimulatory actions on granulocytes. In this report, we show that activated human polymorphonuclear leukocytes (PMN) are able to produce significant amounts of GRO alpha. Lipopolysaccharide (LPS), tumor necrosis factor-alpha (TNF alpha), and yeast particles opsonized with IgG (Y-IgG) had the ability to induce GRO alpha release, with Y-IgG being the most potent stimulus. The extracellular production of GRO alpha was also modulated by both interferon-gamma (IFN gamma) and interleukin-10 (IL-10). IFN gamma significantly inhibited the production of GRO alpha by PMN stimulated for 2 hr with LPS, TNF alpha, or Y-IgG, but potentiated the production of GRO alpha in cells stimulated for 18 hr with LPS and TNF alpha. IL-10 moderately suppressed the Y-IgG-induced production of GRO alpha, but strongly inhibited the action of LPS and potentiated the effect of TNF alpha. As revealed by Northern blot analysis, the extracellular production of GRO alpha under the experimental conditions used did not always correlate with parallel changes at the level GRO alpha mRNA expression, suggesting that production of GRO alpha by PMN might be regulated at post-transcriptional, translational, or post-translational level. These findings identify a novel biological function of PMN, likely involved in the modulation of the acute inflammatory response.

Three intragenic microsatellites of the CFTR gene, a TA and a CA repeats, namely IVS17bTA and IVS17bCA, located in intron 17b and a CA repeat (IVS8CA) located in intron 8 of the CFTR gene, were analyzed in a large sample of Italian cystic fibrosis (CF) and normal chromosomes. Linkage disequilibrium was evaluated between each marker and difference CF mutations on a total of 377 CF and 358 normal chromosomes. Our results are consistent with the hypothesis that all delta F508 chromosomes derive from a single mutational event. The same hypothesis is valid for mutations G542X, N1303K, 1717-1G-->A, which might have been originated more recently than delta F508.

Molecular diagnosis of cystic fibrosis (CF) in the Italian population, based on the detection of the deltaF508 mutation (51.2% of CF chromosomes), provides full informativity for prenatal diagnosis (PDN) in about 28% of families at risk. Identification of the predominant non-deltaF508 mutations allows the characterization of about 70% of CF chromosomes, making approximately 48% of couples fully informative. In families where at least one chromosome remains uncharacterized, allele segregation is still determined using RFLPs closely linked to the CF gene. The recent identification of three polymorphic clusters of dinucleotide repeats (IVS8/GT, IVS17b/TA and IVS17b/CA) led us to evaluate whether their analysis might improve feasibility studies for prenatal diagnosis or heterozygote identification. One hundred nuclear families with a CF child, reflecting the general Italian deltaF508 mutation distribution, were genotyped for the three microsatellites. In this study microsatellite analysis using IVS8/GT and IVS17b/TA allowed the identification of both parental CF chromosomes in 94% of couples; inclusion in the study of the less polymorphic repeat locus, IVS17b/CA, slightly improved this percentage (97%). Hence, a strategy involving primarily the detection of the deltaF508 mutation and secondarily microsatellite analysis makes possible PDN of CF in virtually all Italian CF families.

The delta 508 mutation accounts for about 53% of the molecular defects causing cystic fibrosis (CF) in Italy. The numerous additional mutations detected so far are all relatively rare, and about 30% of CF chromosomes carries unknown mutations in our patients. In order to identify the non-delta F508 mutations causing CF in our population, we performed GC-clamped denaturing gradient gel electrophoresis (DGGE) on 9 exons of the cystic fibrosis transmembrane conductance regulator (CFTR) gene in a sample of 86 Italian CF patients carrying unknown mutations on at least one chromosome. Direct sequencing of 17 samples showing an altered electrophoretic mobility allowed the identification of four new mutations (541delC, R347H, R352Q, and E585X), five mutations already known (G85E, I148T, G178R, 1078delT, and R347P), and one rare variant (1898 + 3A-->G). The strategy based on GC-clamped DGGE represents an efficient and rapid approach for mutation detection for those genetic diseases, such as CF, in which a large number of rare molecular defects has been described.

Grade III placentas were found in 133 (12 per cent) of 1,082 pregnant women evaluated ultrasonographically after 20 weeks of gestation. Associated biparietal diameter measurements ranged from 7.2 to 10 cm. In 11 women with no complications known to accelerate fetal lung maturity or to induce premature placental maturation, grade III placentas were observed between 30 and 33 weeks of gestation. Gestational ages determined according to the growth adjusted sonographic age method of Sabbagha were accurate within one to three days. Considering the risk of hyaline membrane disease at those gestational ages, a grade III placenta cannot be equated with fetal lung maturity. Criteria other than placental grading should be used to assess the pulmonary status of the fetus.

Serum lipid-bound sialic acid (LSA) was measured with a recently described procedure in 108 healthy subjects and in 138 patients with a variety of solid tumors and hematologic malignancies. At the time of serum sampling, 128 patients had active disease and 10 patients had no evidence of disease. LSA was elevated in 104 of 128 (81.2%) patients with active disease, while carcinoembryonic antigen, analyzed in 74, was elevated only in 21 (28.4%) (P less than 0.05). Sensitivity of the serum LSA test ranged from 66% for breast and gastrointestinal cancer to 92% for lung cancer. In patients with lung cancer, ovarian cancer or Hodgkin's disease, LSA was correlated with the extent of disease and it also proved to be useful in following the course of disease. Our preliminary data indicate that this test can be used as a monitor of tumor burden.