Functional aspects of Toll-like receptor/MyD88 signalling during protozoan infection: focus on Toxoplasma gondii.

Toll-like receptor (TLR)/MyD88 signalling has emerged as a major pathway of pathogen recognition in the innate immune system. Here, we review recent data that begin to show how this pathway controls the immune response to protozoan infection, with particular emphasis on the opportunistic pathogen Toxoplasma gondii. The various ways that the parasite activates and suppresses TLR/MyD88 signalling defines several key principals that illuminate the complexities of the host-pathogen interaction. We also speculate how TLR/MyD88 signalling might be exploited to provide protection against Toxoplasma, as well as other protozoa and infection in general.

Dysregulation of macrophage signal transduction by Toxoplasma gondii: past progress and recent advances.

The opportunistic protozoan parasite Toxoplasma gondii is well known as a strong inducer of cell-mediated immunity, largely as a result of proinflammatory cytokine induction during in vivo infection. Yet, during intracellular infection the parasite suppresses signal transduction pathways leading to these proinflammatory responses. The opposing responses are likely to reflect the parasite's need to stimulate immunity allowing host survival and parasite persistence, and at the same time avoiding excessive responses that could result in parasite elimination and host immunopathology. This Review summarizes past and present investigations into the effects of Toxoplasma on host cell signal transduction. These studies reveal insight into the profound suppression of proinflammatory cytokine responses that occurs when the parasite infects macrophages and other cells of innate immunity.

Safety and efficacy results from an international expanded access programme to bortezomib for patients with relapsed and/or refractory multiple myeloma: a subset analysis of the Australian and New Zealand data of 111 patients.

BACKGROUND: Bortezomib has been shown to be a safe and efficacious for the treatment of relapsed and refractory multiple myeloma (MM). Here we report a subset analysis of Australian and New Zealand data from the International Extended Access Programme for bortezomib. METHODS: Patients with more than or equal to two prior lines of therapy were given bortezomib 1.3 mg/m(2) (i.v. bolus days 1, 4, 8, 11) for up to eight 21-day cycles (C). Dexamethasone, 20 mg/day p. o. on the day of, and day after, bortezomib was added after C2 for progressive disease or after C4 for stable disease. Efficacy was assessed using modified Southwest Oncology Group criteria in the intent-to-treat group. Results were compared between the Australian and New Zealand and international cohort. RESULTS: One hundred and eleven patients from 16 centres (55% men, median age 61.9 years) had a median of 5.2 +/- 2.8 treatment cycles of bortezomib. Among them, 82% had > or =3 prior therapies. Grade 3-4 treatment-related adverse events were reported in 57 patients (52%); the most common were thrombocytopenia (25.7%), anaemia (8.3%), peripheral neuropathy (7.3%) and diarrhoea (7.3%). Responses were evaluable in 106 patients: 22% achieved a best response of complete response/response and 20% partial response (overall response rate of 42%). Median times to first and best responses were 42 days and 69 days, respectively. Compared with the international cohort, the cohorts from Australian and New Zealand showed inferior overall response rates (54 vs 42%, P = 0.001), possibly due to heavier pretreatment (82% greater than or equal to three prior therapies vs 68%, P < 0.001). CONCLUSION: Our analysis confirms that bortezomib is safe and effective in relapsed and refractory MM in a real-life clinical setting.

Cotton dust-mediated lung epithelial injury.

To determine if constituents of cotton plants might play a role in byssinosis by injuring pulmonary epithelium, we added extracts of cotton dust, green bract, and field-dried bract to human A549 and rat type II pneumocytes. Injury was measured as pneumocyte lysis and detachment, and inhibition of protein synthesis. Extracts of cotton dust and field-dried bract produced significant dose- and time-dependent lysis and detachment of both target cells, while green bract extract was less damaging. Extracts treated with polyvinylpolypyrrolidone to remove tannins produced significantly less injury. In contrast, purified 5,7,3',4'-tetrahydroxy-flavan 3,4-diol (THF), a tannin in cotton dust and bract, caused substantial cell damage. Field-dried bract extract and THF also produced dose-dependent inhibition of pneumocyte protein synthesis. Endotoxin levels did not correlate with observed injury. THF added to rat tracheal explants caused epithelial disruption and desquamation, whereas endotoxin did not. Instillation of cotton dust and field-dried bract extract in rat lungs produced disruption of bronchial epithelium and smooth muscle constriction, while polyvinylpolypyrrolidone-treated cotton dust extract produced no injury. These findings suggest that extracts of cotton plants are toxic to alveolar, tracheal, and bronchial epithelium and that THF or other tannins may be the responsible agents.

An individualised risk-adapted protocol of pre- and post transplant zoledronic acid reduces bone loss after allogeneic stem cell transplantation: results of a phase II prospective trial.

Bone loss occurs frequently following allogeneic haematopoietic stem cell transplantation (alloSCT). The Australasian Leukaemia and Lymphoma Group conducted a prospective phase II study of pretransplant zoledronic acid (ZA) and individualised post-transplant ZA to prevent bone loss in alloSCT recipients. Patients received ZA 4 mg before conditioning. Administration of post-transplant ZA from days 100 to 365 post alloSCT was determined by a risk-adapted algorithm based on serial bone density assessments and glucocorticoid exposure. Of 82 patients enrolled, 70 were alive and without relapse at day 100. A single pretransplant dose of ZA prevented femoral neck bone loss at day 100 compared with baseline (mean change -2.6±4.6%). Using the risk-adapted protocol, 42 patients received ZA between days 100 and 365 post alloSCT, and this minimised bone loss at day 365 compared with pretransplant levels (mean change -2.9±5.3%). Femoral neck bone loss was significantly reduced in ZA-treated patients compared with historical untreated controls at days 100 and 365. This study demonstrates that a single dose of ZA pre-alloSCT prevents femoral neck bone loss at day 100 post alloSCT, and that a risk-adapted algorithm is able to guide ZA administration from days 100 to 365 post transplant and minimise further bone loss.

The graft-versus-lymphoma effect: clinical review and future opportunities.

Numerous lines of preclinical and clinical evidence support the existence of a graft-versus-leukemia effect, but less evidence supporting a comparable graft-versus-lymphoma effect exists. We review here current clinical data addressing the graft-versus-lymphoma effect, including comparisons of autologous, syngeneic, and allogeneic transplantation; responses to immunomodulation; and responses to nonmyeloablative stem cell transplantation. Despite several limitations of the data, we believe that there is sufficient evidence suggesting a significant graft-versus-lymphoma effect. In addition, we discuss approaches for clinical management of lymphoma patients, opportunities for mechanistic studies afforded by donor leukocyte infusions and nonmyeloablative transplantation, and suggestions for clinical studies to further define the magnitude and applicability of the graft-versus-lymphoma effect.

Use of the second-generation antipsychotic, risperidone, and secondary weight gain are associated with an altered gut microbiota in children.

The atypical antipsychotic risperidone (RSP) is often associated with weight gain and cardiometabolic side effects. The mechanisms for these adverse events are poorly understood and, undoubtedly, multifactorial in etiology. In light of growing evidence implicating the gut microbiome in the host's energy regulation and in xenobiotic metabolism, we hypothesized that RSP treatment would be associated with changes in the gut microbiome in children and adolescents. Thus, the impact of chronic (>12 months) and short-term use of RSP on the gut microbiome of pediatric psychiatrically ill male participants was examined in a cross-sectional and prospective (up to 10 months) design, respectively. Chronic treatment with RSP was associated with an increase in body mass index (BMI) and a significantly lower ratio of Bacteroidetes:Firmicutes as compared with antipsychotic-naïve psychiatric controls (ratio=0.15 vs 1.24, respectively; P<0.05). Furthermore, a longitudinal observation, beginning shortly after onset of RSP treatment, revealed a gradual decrease in the Bacteroidetes:Firmicutes ratio over the ensuing months of treatment, in association with BMI gain. Lastly, metagenomic analyses were performed based on extrapolation from 16S ribosomal RNA data using the software package, Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PICRUSt). Those data indicate that gut microbiota dominating the RSP-treated participants are enriched for pathways that have been implicated in weight gain, such as short-chain fatty acid production.

Larvae-induced plasma membrane wounds and glycoprotein deposition are insufficient for Trichinella spiralis invasion of epithelial cells.

Trichinella spiralis L1 larvae infect susceptible hosts by invading epithelial cells that line the small intestine. Invasion also occurs in vitro when larvae are inoculated into cultures of epithelial cells from several different animal species. To further investigate the mechanism of invasion, we studied the interaction of larvae with the rat epithelial cell line IEC-6. Larvae did not invade IEC-6 cells, but did cause the cells to take up parasite glycoproteins. Glycoprotein bearing cells remained viable and were detectable in monolayers for as long as 24 h, suggesting that the glycoproteins were not toxic for cells. Immunofluorescence revealed that parasite glycoproteins localized in the nuclei, mitochondria and cytoplasm and we found evidence for selection of certain molecules between nuclear and cytoplasmic compartments. Using fluorescent dextrans as fluid phase markers we found 17-38% of the cells in inoculated monolayers were engorged with dextran and that dextran was free in the cytoplasm. Increased dextran uptake was not lethal, required the presence of activated larvae, and was often associated with uptake of parasite glycoproteins. These observations suggest that larvae caused plasma membrane wounds. Our results showed that neither delivery of glycoproteins nor mechanical wounding is sufficient to allow entry of the parasite into resistant epithelial cells. Because both invasion-resistant and susceptible epithelial cells undergo non-lethal wounding, we propose that larvae-induced injury to epithelial cells may result in release of cell-specific mediators that signal larva to invade a particular cell line or, alternatively, to ignore it.

Immunologic responses to inhaled cotton dust.

Byssinosis, a respiratory disease of workers on cotton, flax, and soft hemp, is classically characterized as shortness of breath, cough, and chest tightness on Mondays or the first day of return to work after a time off. Exposure to these vegetable dusts can also result in other respiratory diseases, and the term cotton dust-induced respiratory disease (CDIRD) is introduced. Although clinically characterized for more than a century, the underlying pathogenesis of CDIRD remains obscure. An allergic pathogenesis has been proposed. This article reviews previous and current research findings supporting this mechanism and raises the possibility that, in some individuals, CDIRD may be due to pre-existing or occupationally induced mold allergy.

Respiratory effects in toluene diisocyanate manufacture: a multidisciplinary approach.

A new plant manufacturing toluene diisocyanate (TDI) has provided a unique opportunity to investigate the effects of TDI vapor inhalation on respiratory health in a group of exposed workers who have been studied prior to the start of plant operation. In order to establish dose-response relationships and determine host factors, complete biologic monitoring, including pulmonary function and immunologic studies, has been performed concurrently with a comprehensive environmental monitoring program including continuous sampling for atmospheric concentrations of TDI. Study groups include workers with regular exposure to TDI in production jobs (83), workers with intermittent contact with this vapor, usually in maintenance (28), and a control group of workers employed outside the TDI area (55). This population is being followed for a period of 5 yr. The plant began operations in August 1973 with start-up procedures completed by the end of October. TDI spills occurred for numerous reasons, usually attributed to pump failure and resultant line blockage. Significant exposures also occurred in the drumming operation. The influence of these malfunctions is noted in the continuous monitoring data on atmospheric TDI concentrations which continue to reveal frequent excursions above the threshold limit value (TLV) of 0.02 ppm ceiling. These data are presented in relation to time and plant location. Although the first full year follow-up following initial exposure was not complete, certain preliminary clinical observations were made. A number of workers had episodes of acute respiratory symptoms related to single exposure to an irritant gas at work, usually either TDI or phosgene. It appears that two or three workers in the study population have become "clinically sensitized" to TDI and have been removed from regular TDI exposure. To date, the total number of workers who report the presence of recurring respiratory symptoms has not increased in comparison with the pre-exposure survey. Pulmonary function data after one full year of TDI exposure are not yet available. Pre- and post-shift ventilatory function studies do not indicate significant differences between the exposed and control groups. Selected individuals had carefully controlled inhalation challenge tests to monitored concentrations of TDI vapor under laboratory conditions. In workers suspected of having become "sensitized", immediate and/or late air flow obstruction was demonstrated and could be related to dose of inhaled TDI.

Reactions during work shift among cotton mill workers.

Lung function and blood neutrophils were measured in 13 cotton mill workers on different days of the work week. Antigen was prepared from cotton dust and a radioallergosorbent (RAST) test made on serum and plasma. The exposure was determined as airborne dust and bacterial endotoxin using vertical elutriators and personal samplers. Reductions in FEV1 and increased blood neutrophils were found on Monday at an exposure level of 0.32 to 0.48 mg respirable dust/m3 and 0.19 to 0.28 micrograms endotoxin/m3. The changes in blood neutrophils correlated with endotoxin levels. The FEV1 decreases were present on Monday but not on Wednesday and Friday. The RAST ratios were low and not correlated with FEV1. No systematic changes in RAST ratios were present over the Monday shift. The results suggest that the importance of immunologic reactions other than type 1 should be investigated.

Role of inhalation challenge testing in the diagnosis of isocyanate-induced asthma.

Results of isocyanate challenge tests performed on 63 workers referred with a diagnosis of probable isocyanate asthma between 1974 and 1988 were reviewed. Thirty (48 percent) had an acute episode of asthma with a greater than 20 percent decline in FEV1 following subirritant exposure to isocyanates. No difference in the frequency or type of respiratory complaints between isocyanate reactors and nonreactors was found. No differences in lung function results were present when comparing smoking and ex-smoking reactors and nonreactors. In never-smokers with complaints consistent with isocyanate-induced asthma, the presence of obstructive lung disease increased the likelihood that isocyanate-induced asthma was present. Bronchial responsiveness to methacholine occurred in nearly all isocyanate reactors but predicted isocyanate-induced asthma in only 68 percent of the workers. In nearly all cases of challenge-confirmed toluene diisocyanate (TDI)-induced asthma, a 15-min exposure to 20 ppb of the commercial TDI mixture (80:20 2,4:2,6) provoked asthma. Conversely, in the absence of an asthmatic response following exposure to this dose for this duration, a second exposure at this concentration for a longer time would be reasonable to confirm the absence of isocyanate-induced asthma. Among workers employed in the production of polyurethane foam and confirmed to have TDI-induced asthma by inhalation challenge to the different TDI isomers, there appeared to be increased airway reactivity to the 2,6 isomer. This may have relevance to the frequency and intensity of respiratory symptoms that workers with TDI-induced asthma develop in differing industrial settings.

Absence of hyperresponsiveness to methacholine in a worker with methylene diphenyl diisocyanate (MDI)-induced asthma.

A 29-year-old man had a persuasive history of respiratory illness following exposures to methylene diphenyl diisocyanate (MDI). He was evaluated by measuring bronchial reactivity to methacholine, both before and after controlled laboratory exposures to MDI. Despite evidence of progressive declines in FEV1 with increasing (but subirritant) doses of MDI on three consecutive days, there was no bronchial hyperresponsiveness to methacholine, before or after MDI challenge. We conclude that the absence of nonspecific bronchial hyperresponsiveness does not exclude the possibility of isocyanate asthma. In the face of a compelling history, a negative result of methacholine challenge should not deter observation or laboratory testing for specific respiratory allergy to these chemicals.

Decreased renal excretion of beta-hexosaminidase in adults with insulin-dependent diabetes mellitus and normal renal function.

The activities of three lysosomal hydrolases and creatinine levels were measured in the plasma and urine of 11 adults (mean age, 28.1 years) with insulin-dependent diabetes mellitus and 14 non-diabetic controls (mean age, 27.9 years). All of the patients were free of diabetic complications and non exhibited microalbuminuria. Fractional enzyme excretion (FEE) values between the two groups of subjects were calculated and compared for the following enzymes: beta-hexosaminidase (N-acetyl-glucosaminidase), beta-glucuronidase and alpha-galactosidase. The FEE value was calculated as the ratio of enzyme clearance to creatinine clearance. Relative to the non-diabetic control group, the FEE value for beta-hexosaminidase was approximately 2-fold lower (P = 0.02) in the diabetic subjects (means, 0.424 vs. 0.242, respectively). The FEE values for beta-glucuronidase and alpha-galactosidase were not significantly different (P > 0.4) between the diabetic and control groups. These easily measured biochemical parameters in blood and urine and the resultant FEE value for beta-hexosaminidase may provide a means of assessing subtle deteriorative changes in renal function which occur in the early stage of diabetes before the onset of clinically evident complications.

Use of 4-heptylumbelliferyl-beta-D-glucoside to identify Gaucher's disease heterozygotes.

Three fluorometric beta-glucosidase assays were compared for their ability to identify Gaucher's disease heterozygotes, using leukocytes as the source of enzyme: the pH 5.5-taurocholate assay of Peters et al.; the conduritol B epoxide dependent variation of that assay; and the newly developed method described herein. While the first two procedures utilize the standard substrate 4-methylumbelliferyl-beta-D-glucopyranoside to estimate beta-glucosidase activity, the new assay uses 4-heptylumbelliferyl-beta-D-glucoside as (C7UGlc) substrate. Use of this substrate enhances the specificity of the method for lysosomal glucocerebrosidase, thereby minimizing the contribution of the nonspecific cytosolic beta-glucosidase to estimates of substrate hydrolysis. Using Student's t test for the three assays examined, the C7UGlc assay procedure was determined to have the lowest p value (p less than 0.001) and highest t value (t = 4.95) for the discrimination between the mean glucocerebrosidase value of control and obligate Gaucher heterozygote samples. The high reliability and simplicity of the C7UGlc assay lends adequate reason to favor this assay for regular clinical diagnosis of Gaucher heterozygotes.

The respiratory effects of cotton dust.

It is apparent that occupational exposure to cotton dust, and in vivo exposure to its components, may result in a number of cellular and humoral events. These include specific antibody formation, mediator release, complement activation, endotoxin effects, and related phenomena. The role of these, either singly or in combination, in the pathogenesis of cotton dust-induced lung disease awaits elucidation. Provocative inhalation challenge studies, using well-characterized standard cotton dusts with careful monitoring of physiologic effects, and in vitro assays will provide answers to some of these questions. Development of a suitable animal model would prove to be invaluable for studying the etiology of the disease and might permit evaluation of the relationship of the acute disease to any chronic effects. It is likely that several pathogenetic mechanisms may be found to be operative and that other, as yet untested, effects such as activation of arachidonic acid metabolism may play an important role in the development of this disease.

Sex differences in analgesic response to ibuprofen are influenced by expectancy: a randomized, crossover, balanced placebo-designed study.

AIM: To determine whether there is a sex difference in placebo and ibuprofen analgesia expectancy. METHODS: We measured detection and tolerance thresholds for electrically induced pain in the ear lobe in healthy subjects (10 male, 10 female) to study sex differences in expectancy following either ibuprofen 800 mg or placebo in four different expectancy states. Subjects took ibuprofen or placebo in a two-by-two factorial design (the balanced placebo design). We randomly assigned subjects to start in one of the four expectancy states. We analysed the results using analysis of variance for repeated measures with baseline pain as a covariate. RESULTS AND CONCLUSION: We found no sex difference in baseline pain threshold or tolerance levels. When partitioned by sex and expectancy state, analgesia only occurred in males during positive expectancy states at 2, 3 and 4 h post-placebo, and at 1 and 2 h post-ibuprofen. The time course of analgesic action in males was as expected considering the pharmacokinetic profile of ibuprofen. Our study found that dosages of 800 mg of ibuprofen are ineffective in producing analgesia in women regardless of their expectations. We hypothesize that ibuprofen analgesia is produced by a combination of specific pharmacological effects and a non-specific beta endorphin-mediated placebo effect. Whatever the mechanism responsible for the analgesic response seen in males, this research re-emphasizes the importance of psychological factors in determining drug response. It also shows that these factors can differ between men and women, and thus the contribution of psychological factors on analgesia needs to be seriously re-evaluated.