Idiopathic sudden sensorineural hearing loss and ménière syndrome: The role of cerebral venous drainage.

OBJECTIVES: To evaluate the influence of cerebral venous drainage on the pathogenesis of idiopathic sudden sensorineural hearing loss (ISSHL) and Ménière syndrome (MD). DESIGN: Observational, prospective, cohort study. SETTING: ENT and Cardiology Departments (University of Bari, Policlinico Hospital, Bari, Italy). PARTICIPANTS: We enrolled 59 consecutive patients (32 males, mean age 53.05 + 15.37 years): 40 ISSHL and 19 MD. MAIN OUTCOME MEASURE: All patients underwent physical examination, biochemical evaluation (glycemic and lipid profile, viral serology, C reactive protein, etc), audiometric (tonal, vocal, vestibular evoked myogenic potentials and auditory brainstem response test) and impedentiometric examination. The pure tone average (PTA) was calculated for the following frequencies: 250, 500, 1000, 2000, 3000, 4000, 8000. An echo-color Doppler evaluation of the venous cerebral veins, internal jugular (IJV) and vertebral veins (VV) at supine and 90° position was performed. RESULTS: No morphological alterations were found both in patients and controls. There were no signs of stenosis, blocked flow, membranes, etc. We found lower minimum, mean and maximum velocities in distal IJVs (P = .019; P = .013; P = .022; respectively) and left VVs (P = .027; P = .008; P = .001; respectively) in supine (0°) position in both MD and ISSHL patients as compared to controls. The same was for orthostatic position (90°). We found negative correlations between the velocities in extracranial veins and PTA values: therefore, the worst the audiometric performance of the subjects, the lower the velocities in the venous cerebral drainage. CONCLUSIONS: Idiopathic sudden sensorineural hearing loss and Ménière syndrome patients showed altered venous flow in IJVs and VVs as compared to controls, independently from posture. This different behavior of venous tone control can influence the ear performance and may have a role in the pathogenesis of both diseases.

Goat casein genotypes are associated with milk production traits in the Sarda breed.

The aim of the current work was to analyze, in the Sarda breed goat, genetic polymorphism within the casein genes and to assess their influence on milk traits. Genetic variants at the CSN1S1, CSN2, CSN1S2 and CSN3 gene loci were investigated using PCR-based methods, cloning and sequencing. Strong alleles prevailed at the CSN1S1 gene locus and defective alleles also were revealed. Null alleles were evidenced at each calcium-sensitive gene locus. At the CSN3 gene locus, we observed a prevalence of the CSN3 A and B alleles; the occurrence of rare alleles such as CSN3 B'', C, C', D, E and M; and the CSN3 S allele (GenBank KF644565) described here for the first time in Capra hircus. Statistical analysis showed that all genes, except CSN3, significantly influenced milk traits. The CSN1S1 BB and AB genotypes were associated with the highest percentages of protein (4.41 and 4.40 respectively) and fat (5.26 and 5.34 respectively) (P < 0.001). A relevant finding was that CSN2 and CSN1S2 genotypes affected milk protein content and yield. The polymorphism of the CSN2 gene affected milk protein percentage with the highest values recorded in the CSN2 AA goats (4.35, at P < 0.001). The CSN1S2 AC goats provided the highest fat (51.02 g/day) and protein (41.42 g/day) (P < 0.01) production. This information can be incorporated into selection schemes for the Sarda breed goat.

Evaluation of procalcitonin in hemorrhagic shock: a pilot study.

Abstract: An increased secretion of procalcitonin (PCT) is primarily due to systemic inflammation of bacterial origin, as PCT is used to diagnose and manage sepsis. However, other conditions can induce high plasma levels of PCT, and hemorrhagic shock may be one of these as we found in clinical practice. The aim of this pilot, observational and prospective study was to investigate the role of PCT in hemorrhagic shock and if it could help in distinguishing between different types of shock. We enrolled 15 patients who entered the shock room of our Emergency Department (ED) with a diagnosis of hemodynamic shock, defined as hypotension (systolic blood pressure < 90 mmHg, or medial arterial pressure < 65 mmHg), and/or elevated lactate level (> 2 mmol/L), with one or more signs of cerebral or systemic hypoperfusion. For all the patients we dosed PCT at the time of admission, and we collected them into three different groups - septic, hemorrhagic and mixed shock - based on clinical presentation and laboratory and instrumental examination. First results did not show a significant increase of PCT in patients with hemorrhagic shock alone (average 0.12 ± 0.07 ng/mL), while PCT levels were similarly high in those with septic and mixed shock (17.63 ± 32.16 and 24.62 ± 33.02 respectively). PCT is not a marker of bleeding shock and does not help in distinguishing if bleeding or sepsis have the major impact on hemodynamics in those with mixed shock. However, patients with sepsis usually access the ED a few days after the initial infectious and inflammatory process has begun, while those with a major bleeding ask for intervention at the very first beginning. Thus, it may be helpful to see is PCT levels rise after some time from the bleeding start, or to investigate a different biomarker that rises earlier in course of systemic disfunction, such as presepsin. Finally, we also aimed at investigating if PCT levels would show any correlation with age of patients, regardless of the type of shock: results provided an higher PCT in individuals ≥ 80 years old, than in those < 80 years old.

Management of maxillary artery pseudoaneurysm in Emergency Department: a narrative review.

Abstract: Tooth extraction is a common procedure that is performed routinely and is associated with very few risks. The formation of a pseudoaneurysm as a direct result of tooth extraction has not been widely reported in published studies; it is more frequent as a complication of orthognathic surgery (1). The purpose of this paper is to describe the literature of maxillary artery pseudoaneurysm and its diagnosis and treatment in the Emer-gency Department. The search engine we used is Pubmed. 39 studies were analyzed; mainly, they were case reports. In this study, we will analyze the cases of pseudoaneurysm formation following dental extraction and orthognotia surgery which are reported in literature.

Effect of antigen/antibody ratio on macrophage uptake, processing, and presentation to T cells of antigen complexed with polyclonal antibodies.

Activation of a galactosidase-specific murine T hybridoma clone and of a human tetanus toxoid-specific T clone by antigen-presenting cells (APC) was used to evaluate the regulatory function of antibodies complexed with the relevant antigen. Complexed antigen, in fact, is taken up with high efficiency thanks to Fc receptors borne by APC. Antibody/antigen ratio in the complexes proved to be a critical parameter in enhancing antigen presentation. Complexes in moderate antibody excess provided optimal T cell activation independently of the physical state of the complexes (precipitated by a second antibody or solubilized by complement). Complexes in extreme antibody excess, on the contrary, did not yield T cell activation although taken up by APC efficiently. The effect of antibodies at extreme excess was observed with substimulatory dose of antigen (loss of potentiation) and with optimal dose of antigen (loss of stimulation). An excess of specific polyclonal antibodies hampers proteolytic degradation of antigen in vitro, supporting the view that a similar mechanism may operate within the APC that have internalized immune complexes in extreme antibody excess. The possibility that immune complex forming in extreme antibody excess may turn off the T cell response is proposed as a regulatory mechanism.

Mediterranean diet and the prevention of non-alcoholic fatty liver disease: results from a case-control study.

OBJECTIVE: Few studies report that Mediterranean dietary (MD) pattern has a beneficial role in the progression of non-alcoholic fatty liver disease (NAFLD). Evidence on its potential effect on the onset of disease are, however, scanty. With our study, we evaluated whether MD affects the risk of NAFLD with a large case-control study performed in Italy. PATIENTS AND METHODS: Three hundred and seventy-one cases of NAFLD and 444 controls were questioned on the demographic data and their dietary habits before diagnosis. Additionally, information about lifestyles and other related diseases, such as hypertension and diabetes mellitus were collected. The MD adherence was assessed using a pre-defined Mediterranean Diet Score (MDS). Odds ratios (OR) and 95% confidence intervals (CI) were obtained using a multiple logistic regression model. RESULTS: A high adherence to the MD is significantly associated with decreased risk of NAFLD (OR: 0.83 95% CI: 0.71-0.98). When the different MD components were examined separately, higher legumes consumption (OR: 0.62 95% CI: 0.38-0.99) and high fish consumption (OR 0.38 95% CI: 0.17-0.85) were reported to be protective against NAFLD. CONCLUSIONS: Our study shows that a high adherence to the MD decreases the risk of NAFLD.

Priming of human CD4+ antigen-specific T cells to undergo apoptosis by HIV-infected monocytes. A two-step mechanism involving the gp120 molecule.

The study of the pathology of HIV-1 infection in chimpanzees supports the idea of the crucial role of HIV-infected monocytes in the pathogenesis of AIDS, although viral mechanisms that lead to T cell dysfunction and deletion during HIV infection are still unclear. We show here that HIV-1-infected antigen-presenting monocytes (APCs) are able to prime in vitro non-HIV-infected antigen-specific CD4+ T cell lines or peripheral blood CD4+ T cells to undergo apoptosis after antigen-specific restimulation. The priming of T cells for apoptosis occurs in the absence of HIV replication in the T cells. Priming for apoptosis required two concomitant signals present on the same APC, an antigenic stimulus and a second signal provided by the HIV gp120 protein as demonstrated by the use as APCs of EBV-LCLs infected with different recombinant deleted proviruses or transfected with different HIV proteins. These results provide a mechanism for the priming for apoptosis of T cells in HIV-infected patients, implicating a role for HIV-infected APCs in the induction of T cell dysfunction and depletion in AIDS.

Calorimetric study of concanavalin A binding to saccharides.

The binding of concanavalin A in the dimer form to various saccharides has been studied by calorimetry, and estimates of the binding enthalpy and binding constants have been calculated. Methyl alpha-D-mannoside and methyl alpha-D-glucoside have a -- delta H0 of 21.5 and 11.5 kJ/mol, respectively, at both pH 4 and 4.5. The p-nitrophenyl derivatives react with enthalpic values of 15.6 and 14.6 kJ/mol. The galactosepyranosides show no heat effects during mixing with the protein solutions. The apparent binding enthalpies calculated from the variations of the equilibrium constants with temperature are in good agreement with the values measured experimentally. The two binding sites of the dimer form of concanavalin A are equal and independent, and the low enthalpies obtained do not justify a large conformational change during the reaction. The binding reaction has also been estimated for other sugars normally contained in glycoproteins.

Calorimetric study of manganese binding to concanavalin A.

The reaction of concanavalin A with Mn2+ has been studied calorimetrically. The binding enthalpy was measured at two different temperatures, 25 and 30 degrees C, in 10(-3) M acetate buffer; it was found to be constant between pH 4.0 and 5.0, delta H250 = 95 kJ/mol and delta H300 = 65 kJ/mol, respectively. The two S1 binding sites are identical and independent. Experiments at pH 5.6 are distorted by the heat of aggregation, which is several times higher than the heat of binding. Aggregation was demonstrated by spectrophotometric experiments and by light scattering. The presence of Mn2+ increases the stability of the protein molecule.

Quantitative competitive reverse transcriptase-polymerase chain reaction for BCR-ABL on Philadelphia-negative leukaphereses allows the selection of low-contaminated peripheral blood progenitor cells for autografting in chronic myelogenous leukemia.

The Philadelphia (Ph) translocation t(9;22) results in the creation of the BCR-ABL gene, which is now regarded as central to the mechanism that underlies the chronic phase of chronic myelogenous leukemia (CML). From a clinical point of view, BCR-ABL mRNA detection has become the basis for the study of minimal residual disease in CML, particularly when a complete cytogenetic remission is achieved after interferon-alpha (IFN-alpha) therapy or allogeneic stem cell transplantation. We have recently demonstrated that it is possible to mobilize normal peripheral blood progenitor cells (PBPC) in higher rates if this procedure is performed during the early chronic phase. In an attempt to monitor the leukemic cell content of PBPC collections, we used quantitative-competitive RT-PCR (QC-RT-PCR). Thirty consecutive Philadelphia (Ph) chromosome positive patients were enrolled in this study. After chemotherapy and G-CSF, 14 patients achieved 100% Ph-negative metaphases, nine patients had < or =34% and seven patients >34% leukemic metaphases. A total of 116 collection samples were studied. For each sample, BCR-ABL transcript numbers and BCR-ABL/ABL ratio were evaluated. A highly significant correlation between Ph-positive metaphases and BCR-ABL transcript numbers (r = 0.84, P < 0.0001) or BCR-ABL/ABL ratio (r = 0.86, P < 0.0001) was found. For patients that underwent the procedure in early chronic phase, Ph-negative collections showed different levels of BCR-ABL expression. BCR-ABL transcript numbers varied from a median of 100/microg RNA in the first and second leukaphereses, to 500/microg RNA in the third and fourth leukaphereses, and 1500/microg RNA in the fifth leukapheresis (P = 0.002). BCR-ABL/ABL ratio values showed similar kinetics. We have also demonstrated that there is a correlation between low values in BCR-ABL/ABL ratio (< or =0.01) in the reinfused PBPC and the achievement of cytogenetic remission after autografting (chi2 test, P = 0.01). In conclusion, this study demonstrates that QC-RT-PCR for BCR-ABL is a reliable and helpful method for monitoring residual leukemic load in mobilized PBPC, particularly in Ph-negative collections. Moreover, QC-RT-PCR allows selection of the best available collections for reinfusion into patients after myeloablative therapy.

Calorimetric study of the binding reaction of concanavalin A with immunoglublins.

The thermal effects associated with the binding reaction of concanavalin A and immunoglobulins have been measured by calorimetry. IgG solutions do not generate heat on mixing with concanavalin A, confirming the low reactivity of IgG for the lectin molecule. IgM solutions, on the other hand, show a substantial enthalpic contribution of delta H = -24 +/- 1kJ/site for the binding reaction between the polysaccharide chains of human IgM macroglobulin and concanavalin. A stoichiometry is 10 monovalent concanavalin A molecules per intact macroglobulin and two bivalent concanavalin A molecules per two sites on the heavy chain of the reduced macroglobulin subunit.

Calorimetric study of rheumatoid factor binding to human IgG.

The binding of human IgM monoclonal rheumatoid factor (RF) to native human IgG was studied in a flow calorimeter. Thermal titration gave a enthalpy of binding of 5.5 X 10(4) J/mole site for the reaction, with 1 mole of RF binding to 2.5 moles of IgG; this corresponds to one RF subunit per IgG heavy chain. Similar experiments performed with heat-aggregated human IgG gave almost identical results. If the IgG molecule undergoes a conformational change upon heating, this is not such as to affect the enthalpic contribution of binding.

Cytogenetic response to autografting in chronic myelogenous leukemia correlates with the amount of BCR-ABL positive cells in the graft.

OBJECTIVE: An important step in successful autografting of patients with chronic myelogenous leukemia is the delivery of a leukemia-free graft. We conducted this study to determine whether the cytogenetic response after autografting was correlated with the number of BCR ABL-positive cells present within the stem cell grafts. MATERIALS AND METHODS: By BCR-ABL mRNA quantification, we studied the serial pheresis products from 40 Philadelphia (Ph)-positive patients who received ICE/mini-ICE mobilization therapy and underwent autologous stem cell transplantation. We correlated the residual disease within the graft reinfused with the cytogenetic response following transplantation, taking into consideration those responses that lasted 12 months or more. RESULTS: Thirty-two patients received a graft with 0-35% Ph-metaphases and 19 received a graft with BCR-ABL/ABL ratio < or =0.01. After a median of 27 months (range, 12-50) from transplant, 18 patients achieved complete or major cytogenetic response lasting at least 12 months, and 14 of them (78%) received a graft with BCR-ABL/ABL ratio < or =0.01 (range, 0.0003-0.01). Twenty-two patients experienced short-lived responses or had >35% Ph-positive cells in the marrow after transplant, but only 5 of them (23%) had a graft with BCR-ABL/ABL ratio < or =0.01 (range, 0.001-0.01). Therefore, we found a strong association between a BCR-ABL/ABL ratio less than or =0.01 and the achievement of complete or major cytogenetic remission after autografting (chi(2) test, p = 0.0001). Patients reinfused with grafts contaminated at low levels with leukemic cells also showed a longer duration of the response (log-rank test, p = 0.0009). Eleven patients were reinfused with the lowest level of contaminated stem cell collections, according to the BCR-ABL/ABL ratio. None of these patients experienced prolonged neutropenia or thrombocytopenia following stem cell reinfusion and nine of them had long-lasting complete or major cytogenetic responses after transplant. CONCLUSION: This study demonstrates that the number of BCR-ABL positive cells present in a stem cell graft is an important predictive factor for the achievement and the duration of cytogenetic response after autografting. [corrected]

Dendritic cells are potent antigen-presenting cells for in vitro induction of primary human CD4+ T-cell lines specific for HIV gp120.

Activation of T helper cells specific for viral antigens is critical for antibody production and for generation of cytotoxic cells during the immune response to HIV. Since T-cell activation depends on antigen-presenting cells (APCs), it is important to define the cells that have a role in presentation of HIV antigens in general and of gp120 in particular. Peripheral blood mononuclear cells (PBMCs), adherent monocytes (AMs), dendritic cells (DCs) and Epstein Barr virus-transformed B-cell lines (LCLs) were tested for the capacity to present gp120 to a specific T-cell clone. DCs proved to be the most effective APC. Primary T-cell lines were generated from uninfected and unprimed individuals by using different APCs in the presence of gp120 or an immunodominant peptide. T-cell lines specific for gp120 were obtained with PBMCs or DCs as APCs, but not with AMs or LCLs. The data showed that (a) DCs are the most effective APCs for presentation of gp120 to specific T cells and (b) DCs are necessary for in vitro induction of primary T-cell lines specific for gp120.

A simple procedure for freezing and storing lymphocyte panels in trays.

Lymphocytes from different donors may be frozen in Terasaki trays by a simple procedure. This method provides the tissue typing laboratory with a readily available panel that can be routinely used to test for lymphocytotoxic antibodies sera from patients waiting for kidney transplant, transplanted patients and polytransfused individuals. The advantage of this procedure is that expensive programmed freezing apparatus is not required.

An enzymatically active antigen-antibody probe to measure circulating immune complexes. II. E. coli beta-galactosidase in the probe and C1q as the recognition unit.

An enzymatically active probe (beta-galactosidase-anti-beta-galactosidase complex) is used to measure circulating immune complexes (CIC), in a competition assay where probe and CIC are confronted with a 'recognition unit'. The latter is bovine conglutinin in the original description of this method. Here we describe a version utilizing human or bovine C1q. The two techniques are compared for their sensitivity and specificity, on both in vitro formed tetanus toxoid-anti-toxoid complexes and on sera from patients with selected diseases. The results confirm that the two recognition units are sensitive to families of CIC that only partially overlap. The parallel use of conglutinin and C1q yields both quantitative and qualitative information on the nature of CIC in individual sera.

Inhibition of the accessory function of murine macrophages in vitro by cyclosporine.

The accessory function of macrophages, which is strictly related to the induction of T cell activation, has been studied to determine whether it is affected by cyclosporine (CsA). Irradiated spleen cells, used as a source of macrophages, were pulsed overnight with beta-galactosidase (GZ) in the presence of CsA. After washing of the pulsed macrophages, cells from a GZ-specific T cell line were added to cultures and 3H-thymidine incorporation was measured 72 hr later. We found that 500 ng/ml CsA present during macrophage pulsing with GZ reduced T cell proliferation to 5%. On the other hand, 100 ng/ml CsA almost completely abrogated the proliferative response when present for the duration of the culture. Similar results were also obtained using antigen-pulsed peritoneal-adherent macrophages to stimulate the T cell line to proliferate, or a T hybridoma clone to produce interleukin-2 (IL-2). The possibility that CsA actually affects interleukin-1 (IL-1) production by macrophages by inhibiting uninvolved T cells could be ruled out. We conclude that CsA-induced inhibition of T cell functions (proliferation and IL-2 production) is partially due to the effect of the drug on the accessory function of macrophages. This immunosuppressive mechanism of action of CsA on macrophages has not been previously described.

Effect of cyclosporine on the antigen-presenting function of human and murine accessory cells.

Recent reports have challenged the belief that accessory cells are resistant to cyclosporine. Such a tenet was based on the observation that several functions of accessory cells, such as IL-1 production and phagocytosis, are resistant to the drug. On the other hand, when a less primitive, more refined function of accessory cells was examined--i.e., the capacity to take up, process, and present antigen in an MHC-restricted fashion to antigen-specific T lymphocytes, CsA proved to be an effective inhibitor. In contrast to this finding, when antigen was provided in the form of an immune complex prepared with a monoclonal antibody, uptake of antigen--likely mediated by the Fc receptors--and subsequent processing and presentation were not affected by CsA. These results suggest that, depending on whether the antigen is taken up by constitutive or by receptor-mediated endocytosis, accessory cells can be functionally defined as resistant or sensitive to CsA.

Difference in sensitivity to cyclosporine in vitro of human alloreactive lines and clones.

The effectiveness of cyclosporine (CsA) as immunosuppressive agent in human kidney graft rejection is well established. However, in spite of efforts to maintain optimal plasma levels, a fraction of transplanted patients undergo rejection episodes and/or irreversible chronic rejection. This suggests that immunosuppression by CsA cannot control the alloreactive response if there is a high degree of histoincompatibility for HLA or non-HLA antigens, or it has little effect on the "high responder" patient. Both possibilities are difficult to test in the human system. A third hypothesis, the existence of individual CsA resistance, was tested by evaluating the in vitro inhibitory activity of CsA on alloreactive T cell lines from several individuals. A different degree of in vitro sensitivity to the drug was observed among alloreactive lines generated from different individuals and among clones obtained from the same bulk line. The variability at the individual level and at the clonal level may account for the onset of CsA-resistant rejection assuming that in vivo a positive selection in the presence of the drug occurs and allows for the resistant clones, if present, to dominate the sensitive ones.

Allospecific rat-mouse T hybridoma clones generated from kidney graft-infiltrating cells.

In order to dissect the cellular mechanisms that lead to allograft rejection, infiltrating cells can be isolated and expanded in vitro for functional assays. Since the maintenance of these lines and clones is time-consuming, and large numbers of specific cells cannot be easily obtained, we fused the graft infiltrating cells, after in vitro specific expansion, with the murine T lymphoma BW5147. The rat-mouse TT hybridomas thus generated were screened for antigen-dependent interleukin 2 production, for antigen-dependent polyclonal helper activity, and for surface phenotype. The high frequency of specific clones obtained indicates that this is a convenient approach to generate alloreactive hybridoma clones with specific functions from the inflammatory infiltrates of rejected grafts.