Study on hydrophilicity and degradability of chitosan/polylactide-co-polycaprolactone nanofibre blend electrospun membrane.

Electrospinning is an interesting technique to produce polymer membranes made of entangled nanofibres. The technique is raising interest in pharmaceutical and biomedical areas. Either electrospun membranes are studied for tissue regeneration purposes, or incorporation of nanoparticles in electrospun membranes can be an opportunity to control the delivery of drug or to obtain dual drug delivery system. In this work suspensions of hydrochloride chitosan salt in copolymer polylactide-co-polycaprolactone (PLA-PCL) solution were electrospun in order to assess an advanced study for developing polymer nanofibre blend membrane loaded with chitosan polymer. The aim of the work was to investigate the properties and stability of chitosan/PLA-PCL electrospun membranes considering their application for tissue regeneration and drug delivery. The electrospun membranes were characterized for their physico-chemical (FT-IR) morphology (SEM) and in vitro biological properties (cytocompatibility and cells engraftment). Results show that homogeneous electrospun PLA-PCL/chitosan blend nanofibres in the range size 800 nm were obtained. Chitosan was loaded inside the nanofibres up to 27.2% (w/w) without modifying nanofibre shape, and only 6% of the loaded chitosan resulted to be on the nanofibre surface. The presence of chitosan in the nanofibres has shown to accelerate the electrospun membranes degradation in vitro.

Emerging and re-emerging infectious disease in otorhinolaryngology.

SUMMARY: Emerging and re-emerging infectious disease in otorhinolaryngology (ENT) are an area of growing epidemiological and clinical interest. The aim of this section is to comprehensively report on the epidemiology of key infectious disease in otorhinolaryngology, reporting on their burden at the national and international level, expanding of the need of promoting and implementing preventive interventions, and the rationale of applying evidence-based, effective and cost- effective diagnostic, curative and preventive approaches. In particular, we focus on i) ENT viral infections (HIV, Epstein-Barr virus, Human Papilloma virus), retrieving the available evidence on their oncogenic potential; ii) typical and atypical mycobacteria infections; iii) non-specific granulomatous lymphadenopathy; iv) emerging paediatric ENT infectious diseases and the prevention of their complications; v) the growing burden of antimicrobial resistance in ENT and the strategies for its control in different clinical settings. We conclude by outlining knowledge gaps and action needed in ENT infectious diseases research and clinical practice and we make references to economic analysis in the field of ENT infectious diseases prevention and care.

A new solid phase immunoradiometric assay for antithyroid microsomal antibody.

A new sensitive, quantitative, and specific immunoradiometric assay (IRMA) for antithyroid microsomal (anti-M) antibody has been developed. Samples to be tested are incubated within wells of polyvinyl microtiter plates coated with solubilized thyroid microsomal antigen. After removal of unbound material, anti-M antibody is detected by adding purified [125I]antihuman immunoglobulin G (IgG) antibody. Using 1.0 microliter serum, anti-M antibody was found by IRMA in all of the patients with Hashimoto's thyroiditis or idiopathic myxedema (n = 19), in 86% of those with Graves' disease (n = 42), in 10.9% of subjects with other nonautoimmune thyroid disorders (n = 37), and in 8.4% of normal controls (n = 71). A good correlation was found with the results obtained in anti-M antibody tests by passive hemagglutination. Using larger volumes of serum (up to 100 microliters), anti-M antibody detectable by IRMA was found in some patients with Graves' disease and negative passive hemagglutination tests. Quantitative measurements of anti-M antibody by IRMA could be performed using a standard IgG preparation containing high levels of anti-M antibody. The minimal detectable amount ranged between 1-2 ng IgG, corresponding to a sensitivity 15-30 times greater than that of the competitive binding radioassay. We suggest that the present IRMA may be proposed as a general technique for the detection of different organ-specific autoantibodies.

Surface markers and function of circulating thyroid autoantibody-producing cells.

The in vitro synthesis of antithyroglobulin (anti-Tg) and antithyroid microsomal (anti-M) autoantibodies by peripheral blood mononuclear cells (MNC) from patients with autoimmune thyroid diseases was investigated using sensitive immunoradiometric assays. Cultures were carried out in the presence or in the absence of pokeweed mitogen (PWM). Thyroid autoantibodies were undetectable in supernatants of MNC cultures from 9 normal subjects. Supernatants of MNC cultured without PWM had detectable levels of anti-Tg and anti-M in 5 (19.3%) and in 2 (7.7%) of 26 patients with autoimmune thyroid diseases, respectively. In the presence of PWM, a marked increment in the antibody concentrations occurred in all but 1 of these cultures, and the number of positive cultures increased to 13 (50.1%) for anti-Tg and to 15 (57.7%) for anti-M. Studies of MNC fractions depleted of T lymphocytes (non-T cells) were carried out on selected patients showing antibody synthesis only after PWM stimulation. Autoantibody production was not found with non-T cells, but the effect of the mitogen was restored by readdition of T cells. Irradiation (1000 rad) of T cells before coculturing significantly enhanced autoantibody production. With this model no significant functional difference was found between autologous and allogenic T cells from thyroid autoimmune disease patients or from normal subjects. The cells involved in PWM-driven thyroid autoantibody synthesis, as defined by depletion studies, were lymphocytes bearing DR antigens and surface immunoglobulin G (IgG) without detectable surface immunoglobulin M (IgM). Depletion from MNC suspensions of Tg-binding cells abolished PWM-stimulated anti-Tg production, but did not alter the synthesis of anti-M. Further studies were carried out on MNC from a single patient with Hashimoto's thyroiditis, whose non-T cells consistently produced large amounts of anti-M and total IgG in the absence of PWM. The addition of PWM to these unfractionated MNC slightly increased the production of anti-M, but inhibited antibody synthesis after depletion of T lymphocytes. Interestingly, the addition of autologous T lymphocytes to non-T cells inhibited the spontaneous synthesis of anti-M. These data indicate that in vitro synthesis of anti-Tg and anti-M by MNC may be frequently induced by stimulation with PWM in patients with thyroid autoimmune disorders. PWM-stimulated synthesis of thyroid autoantibodies appears to be T-cell dependent and modulated by radiosensitive T lymphocytes. The cells responsible for PWM-dependent thyroid autoantibody synthesis are B lymphocytes with surface membrane IgG and have receptors specific for the autoantigen.(ABSTRACT TRUNCATED AT 400 WORDS)

Lactoferrin inhibits herpes simplex virus type 1 adsorption to Vero cells.

This paper describes the ability of human and bovine lactoferrins (HLf; BLf), iron-binding proteins belonging to the non-immune defense system, to interfere with herpes simplex virus type 1 (HSV-1) infection. Since lactoferrins are known to bind to heparan sulphate proteoglycans and to low density lipoprotein receptor, which in turn act as binding sites for the initial interaction of HSV-1 with host cells, we tested the effect of these proteins on HSV-1 multiplication in Vero cells. Both HLf and BLf are found to be potent inhibitors of HSV-1 infection, the concentrations required to inhibit the vital cytopathic effect in Vero cells by 50% being 1.41 microM and 0.12 microM, respectively. HLf and BLf exerted their activity through the inhibition of adsorption of virions to the cells independently of their iron withholding property showing similar activity in the apo- and iron-saturated form. The binding of [35S]methionine-labelled HSV-1 particles to Vero cells was strongly inhibited when BLf was added during the attachment step. BLf interacts with both Vero cell surfaces and HSV-1 particles, suggesting that the hindrance of cellular receptors and/or of viral attachment proteins may be involved in its antiviral mechanism.

Effect of HSV-2 infection on the expression of HPV 16 genes in CaSki cells.

Human papillomaviruses (HPVs) have been proposed to be the most important etiological factors for cervical cancer although different agents may act in conjunction. Herpes simplex virus type 2 (HSV-2) infection is considered as a possible cofactor to malignant transformation. To examine the influence of HSV-2 infection on the HPV genes expression, CaSki cells bearing 60 to 600 copies of HPV-16 DNA per cell were used as a model system. Twenty hours post HSV-2 infection the mRNA transcripts for HPV-16 early (E1, E2 and E6) and late (L1) genes were analysed by RT-PCR assay. Results indicated that the level of transcription of E1, E2 and E6 genes was up to 3-fold enhanced in HSV-2 infected CaSki cells suggesting that HSV-2 infection could increase the risk of cervical cancer by overexpression of both HPV regulatory and oncogenic genes.

Inhibition of herpes simplex virus infection by negatively charged and neutral carbohydrate polymers.

Different natural and semisynthetic polysaccharides were evaluated for their inhibitory effect on in vitro replication of herpes simplex virus (HSV) types 1 and 2. Some neutral and negatively charged carbohydrates were able to inhibit viral infection by interfering mainly with the adsorption process showing a dose-dependent relationship. Their effect was shown within the concentration range of 200-0.8 micrograms/ml, and the inhibiting compounds were in order of action: dextran sulfate = scleroglucan = lambda carrageenan > glyloid sulfate 4324 > locust beam gum towards HSV-1 and dextran sulfate = glyloid sulfate 4324 = lambda carrageenan > scleroglucan > glycogen sulfate 4435 towards HSV-2. The data obtained indicate that the antiviral activity of polysaccharides was not only related to their electric charge. Other characteristics of the molecules such as the polymeric backbone, the carbohydrate moieties and the degree of polymerization could play a role in influencing their antiviral properties.

Simultaneous detection of HPV and other sexually transmitted agents in chronic urethritis.

BACKGROUND: Many pathogens may be responsible of Non Gonococcal Urethritis (NGU) with the possible occurrence of symptomatic and asymptomatic mixed viral and bacterial infections. In particular, genital papillomaviruses (HPVs) have been searched since they are linked to both benign and malignant lesions of the penis and urethra and the presence of a potential male carried state has received limited scrutiny while the screening of sexually active females has received substantial attention. METHODS: In male patients affected by chronic NGU, the presence of DNA of Chlamydia trachomatis, herpes simplex virus (HSV) type 1 and 2 and human papillomaviruses by PCR and the occurrence of Gram positive and Gram negative micro-organisms, of Mycoplasma hominis and Ureaplasma urealyticum, by conventional cultural methods have been investigated. RESULTS: Results obtained indicated a high percentage of mixed infections, up to 36%. Genital HPV DNA was detected in 31% of specimens positive for two or more agents, and HSV DNA was detected in 10% of studied population. CONCLUSIONS: The concomitant presence of different infectious agents could determine latent, sub-clinical or chronic infections with periodic reactivation. In particular results suggest that HPV and HSV may stimulate cytokine production which can up regulate the expression of other infectious agents and may be responsible for latent chlamydial infections characterised by the persistence of this micro-organism in an altered form, viable but in a culture negative state. Therefore an increased awareness of mixed infections is relevant to define the management and treatment of chronic urethritis.

In vitro study of a double infection by herpes simplex virus type 2 and Chlamydia trachomatis.

Sexually transmitted diseases (STDs) may recognize multiple etiological agents. Among them, Chlamydia trachomatis (Ct) and herpes simplex virus type 2 (HSV-2) cause symptomatic, subclinical and asymptomatic infections of the urogenital tract which can lead to serious sequelae. In the present study the coinfection and superinfection by Ct and HSV-2 in epithelial cultured cells from human cervix (HeLa 229) are described. A double infection, followed by the intracellular synthesis of chlamydial and viral antigens, was established. Both synergistic and interfering phenomena were recorded: viral antigen synthesis resulted increased whereas Ct inclusion bodies were produced to a lower extent.

Detection of viral and bacterial infections in women with normal and abnormal colposcopy.

Signs and symptoms of sexually-transmitted diseases (STD) do not allow any etiological diagnosis in women. Colposcopic findings are seldom pathognomic. Consequently, the microbiology laboratory with the recent availability of molecular diagnostic tools is required to detect the infectious bacterial and/or viral agents involved in STD. In cervical samples of women submitted to gynaecological screening for past or present signs and symptoms of inflammation and with different colposcopic findings, we searched by molecular approaches Chlamydia trachomatis, Mycoplasma genitalium, herpes simplex virus type 1 and 2, adenovirus and 45 genotypes of papillomaviruses and, by cultural methods Mycoplasma hominis and Ureaplasma urealyticum. Colposcopy permitted us to divide the studied population into three groups: 48 women had negative colposcopic findings, 50 presented signs of flogosis and 100 resulted positive for an abnormal transformation zone (ANTZ) and/or for HPV colposcopic findings. Results obtained by microbiological assays indicated that the prevalence of infectious agents did not always correlate with colposcopy. Double and triple infections were found in groups 2 and 3, with mycoplasmas being the most common microrganisms present in association and quite almost copresent with papillomaviruses.

Staff counseling specialist. Responding sensitively to workers.

Due to changes in the health care environment that have expanded their role, nurse administrators have less opportunity to respond adequately to employee concerns. The position of staff counseling specialist was designed to assist nurse administrators at the University of Minnesota Hospitals and Clinics meet the increased demands of their staff. The staff counseling specialist provides a means of narrowing the gap between the staff and nurse executives by addressing employee concerns such as inadequate leadership, communication breakdown, dissonant physician-nurse relations, and other matters adversely affecting the work climate.

A new solid-phase immunoradiometric assay for anti-thyroglobulin autoantibody.

A newly developed sensitive and quantitative immunoradiometric assay (IRMA) for anti-thyroglobulin (anti-Tg) autoantibody is described. Serum samples to be tested are added to wells of polyvinyl microtiter plates coated with human thyroglobulin. After removal of the unbound material, anti-Tg antibody is determined by adding purified 125I-anti-human immunoglobulin G antibody. Using 1.0 microliter of serum anti-Tg antibody was detected in 81.2% of patients with Hashimoto's thyroiditis or idiopathic myxedema (n = 32), in 46.4% of those with Graves' disease (n = 28), in 11.9% of subjects with other thyroid disorders (n = 42) and in 4.2% of normal controls (n = 71). Similar percentages of positive tests were observed by passive hemagglutination (PH) and a good correlation was found between the antibody levels determined by the two techniques. Using larger amounts of serum (100 microliters) detectable anti-Tg antibody by IRMA was found in the majority of patients with thyroid autoimmune disorders who had negative PH tests. Quantitative measurements of anti-Tg antibody by IRMA could be obtained by using purified anti-Tg antibody as standard reference. The minimum detectable amount of anti-Tg antibody was 0.5 ng. The present method is proposed as a simple and convenient technique for quantitative measurement of any antibody, using wells coated with the appropriate antigen.

Skin necrosis due to intravenous heparin.

A patient on intravenous heparin is described who developed an acute hemorrhagic necrosis of her legs. Thrombi without vasculitis were seen in the dermal blood vessels. This appears to be the 4th reported case of skin necrosis after intravenous heparin at sites unrelated to injections. Skin necrosis after intravenous heparin should warn the dermatologist of a possible fatal outcome from myocardial or cerebral infarction.

Antiviral effect of a polysaccharide from Sclerotium glucanicum towards herpes simplex virus type 1 infection.

Among different neutral polysaccharides from natural sources, scleroglucan from Sclerotium glucanicum significantly inhibits the replication of herpes simplex virus type 1 on Vero cells. Scleroglucan belongs to a class of exopolymers, expressed by members of genus Sclerotium and consists of a linear beta-1,3-linked glucopyranose with side chains of single glucopyranose residues linked through beta-1,6 glycosidic bonds. The effective antiviral concentration of this polysaccharide is far from the cytotoxicity threshold and consequently this natural product possesses a good selectivity index. Results obtained in experiments carried out in order to clarify the mechanism of action of this carbohydrate indicate that the block of infection occurs during the very early phases of the viral mutliplication cycle since the highest inhibitory effect took place when it was added during the attachment step. The antiviral effect of scleroglucan seems to be related to its binding with membrane glycoproteins of HSV-1 particles which impedes the complex interactions of the virus with the cell plasma membrane.

Involvement of herpes simplex type 2 in modulation of gene expression of human papillomavirus type 18.

Human papillomaviruses (HPVs) and herpes simplex virus type 2 (HSV-2) can establish latent or persistent infections in the host, and are involved in the aetiology of benign and/or malignant lesions of the urogenital tract. To investigate the putative interaction between these DNA viruses when a double infection occurs, we have studied the effect of HSV-2 infection in HeLa 229 cells containing 10-50 copies of HPV type 18 genomic DNA. Twenty hours post HSV-2 infection, the analysis of mRNA transcripts from E1, E2, E6 early and L1 late HPV18 genes was performed in HeLa cells by a semi-quantitative RT-PCR assay. A modulation of HPV18 E1 and E6 early genes was observed, resulting in a 9-fold and 3-fold increased transcription respectively.

Metal complexes of bovine lactoferrin inhibit in vitro replication of herpes simplex virus type 1 and 2.

The inhibitory effect of bovine lactoferrin (BLf) saturated with ferric, manganese or zinc ions, on the infection of Vero cells by human herpes simplex virus type 1 (HSV1) and 2 (HSV2) was investigated. Viral infectivity determined by intracellular antigen synthesis and plaque formation was efficiently inhibited by metal saturated lactoferrins in a dose-dependent manner. Effective BLf concentrations which reduced the infection by 50% ranged from 5.2 to 31 micrograms ml-1 and were far below the cytotoxicity threshold. Fe3+BLf and Mn2+BLf exhibited selectivity indexes higher than Zn2+BLf and apoBLf for both viruses and the effect was mainly directed towards the early steps of infection. The slight viral inhibition shown by the citrate complexes of the different metals could indicate that the antiviral effect was not significantly influenced by Fe3+, Mn2+ or Zn2+ ions delivered by BLf into the cells.

Superinfection by Listeria monocytogenes of cultured human enterocyte-like cells infected with poliovirus or rotavirus.

A mixed infection with either rotavirus or poliovirus and Listeria monocytogenes was analysed in Caco-2 cells, a tumour-derived cell line, highly susceptible to these pathogens. The multiplication of these pathogens, whose usual site of entry and/or replication is the intestine, was also followed by electron microscopy. Results obtained showed an increase of L. monocytogenes internalisation in cells infected with rotavirus, whereas the preinfection with poliovirus had only a slight interfering effect on bacterial entry. Analysis of L. monocytogenes multiplication in virus-infected cells revealed that rotavirus also promoted bacterial replication, which poliovirus hampered replication. Concerning the effect of Caco-2 cell invasion by L. monocytogenes on viral replication, we observed an increase in rotavirus antigen synthesis but no significant effect on poliovirus yield under our experimental conditions.

In vitro effect of natural and semi-synthetic carbohydrate polymers on Chlamydia trachomatis infection.

The effect of different natural and semi-synthetic polysaccharides on Chlamydia trachomatis multiplication in Hela 229 cells was evaluated. Some neutral, negatively and positively charged carbohydrates were able, in a dose-dependent fashion, to inhibit chlamydial infection by interfering mainly with the adsorption process. The inhibiting compounds, whose effect was shown within the concentration range of 8-200 micrograms/ml, were in order of action: dextran sulphate > glyloid sulphate 4327 > glycogen sulphate 4427 > arabic gum = glyloid > chitosan > glycogen. Data obtained suggested that antichlamydial activity was not only related to the electric charge of these molecules but could also be attributed to other features of their polymeric backbone. Since carbohydrate polymers have also been shown to inhibit the early stages of infection by viral agents causing sexually transmitted diseases, the employment of these molecules for prevention or treatment of mixed viral-C. trachomatis infections can be hypothesized.