Long-term relationships between screening rates, breast cancer characteristics, and overdiagnosis in US counties, 1975-2009.

Effects of mammography screening in the general population are disputed. Screening rates differ greatly between US counties, providing a natural opportunity to investigate effects of screening. We compared mammography screening rates with the types and outcomes of breast cancers diagnosed in US counties. The county screening rate was defined as the proportion of women age ≥40 with ≥1 mammogram in the past 2 years (range, 34-91%). Two periods were analyzed: 1975-2009 (612,941 breast cancer cases, 195 counties) and 1996-2009 (645,057 cases, 211-547 counties). Multiple signs of overdiagnosis were observed: First, breast cancer incidence increased as screening became common. Second, incidence stopped increasing once screening rates stabilized. Third, the increases in incidence were limited to age groups receiving screening. Fourth, the increases were larger in counties where screening became more common. Fifth, the increases were limited to small and early-stage breast cancers (which are consistent with overdiagnosis). Sixth, compensatory reductions in large and advanced-stage breast cancers were much smaller than the increases. Difference-in-differences regression analysis suggested 31% (95% CI: 28-34%) of breast cancers diagnosed in 1996-2009 were overdiagnosed. Screening rates correlated with increased incidence for all hormone receptor statuses, HER2 statuses, and grades. Reductions in breast cancer mortality during 1975-2009 were similar in screened and unscreened age groups. Overall, we found repeated signs that breast cancer overdiagnosis is widespread in the US, but the biological nature of overdiagnosed tumors remains unclear. Mortality benefits of screening, though they may be present and substantial, could not be detected at the population level.

A national UK audit of suprapubic catheter insertion practice and rate of bowel injury with comparison to a systematic review and meta-analysis of available research.

OBJECTIVES: Limited data exist on the risks of complications associated with a suprapubic catheter (SPC) insertion. Bowel injury (BI) is a well-recognized albeit uncommon complication. Guidelines on the insertion of SPC have been developed by the British Association of Urological Surgeons, but there remains little evidence regarding the incidence of this complication. This study uses contemporary UK data to assess the incidence of SPC insertion and the rate of BI and compares to a meta-analysis of available papers. METHODS: National Hospital Episodes Statistics data were searched on all SPC insertions over an 18-month period for operating procedure codes, Code M38.2 (cystostomy and insertion of a suprapubic tube into bladder). Patients age, 30-day readmission rates, 30-day mortality rate, and catheter specific complication rate were collected. To estimate the BI rate, we searched patients who had undergone any laparotomy or bowel operation within 30 days of SPC insertion. Trusts were contacted directly and directed to ascertain whether there was SPC-related BI. PubMed search to identify papers reporting on SPC related BI was performed for meta-analysis RESULTS: 11 473 SPC insertions took place in the UK in this time period. One hundred forty-one cases had laparotomy within 30 days. Responses from 114 of these cases reported one BI related to SPC insertion. Meta-analysis showed an overall BI rate of 11/1490 (0.7%). CONCLUSIONS: This is the largest dataset reported on SPC insertions showing a lower than previously reported rate of BI. We recommend clinicians use a risk of BI of less than 0.25% when counseling low-risk patients.

Glutamine and hyperammonemic crises in patients with urea cycle disorders.

UNLABELLED: Blood ammonia and glutamine levels are used as biomarkers of control in patients with urea cycle disorders (UCDs). This study was undertaken to evaluate glutamine variability and utility as a predictor of hyperammonemic crises (HACs) in UCD patients. METHODS: The relationships between glutamine and ammonia levels and the incidence and timing of HACs were evaluated in over 100 adult and pediatric UCD patients who participated in clinical trials of glycerol phenylbutyrate. RESULTS: The median (range) intra-subject 24-hour coefficient of variation for glutamine was 15% (8-29%) as compared with 56% (28%-154%) for ammonia, and the correlation coefficient between glutamine and concurrent ammonia levels varied from 0.17 to 0.29. Patients with baseline (fasting) glutamine values >900 µmol/L had higher baseline ammonia levels (mean [SD]: 39.6 [26.2]µmol/L) than patients with baseline glutamine ≤ 900 µmol/L (26.6 [18.0]µmol/L). Glutamine values >900 µmol/L during the study were associated with an approximately 2-fold higher HAC risk (odds ratio [OR]=1.98; p=0.173). However, glutamine lost predictive significance (OR=1.47; p=0.439) when concomitant ammonia was taken into account, whereas the predictive value of baseline ammonia ≥ 1.0 upper limit of normal (ULN) was highly statistically significant (OR=4.96; p=0.013). There was no significant effect of glutamine >900 µmol/L on time to first HAC crisis (hazard ratio [HR]=1.14; p=0.813), but there was a significant effect of baseline ammonia ≥ 1.0 ULN (HR=4.62; p=0.0011). CONCLUSIONS: The findings in this UCD population suggest that glutamine is a weaker predictor of HACs than ammonia and that the utility of the predictive value of glutamine will need to take into account concurrent ammonia levels.

A study examining the relationship between alexithymia and challenging behaviour in adults with intellectual disability.

BACKGROUND: Research suggesting that people with intellectual disabilities (ID) have difficulties in recognising emotions provides a rationale for studying alexithymia in this population. A number of studies have found a relationship between alexithymia and challenging behaviours in various populations and this study aims to discover if this is the case for people with ID. METHOD: Cross-sectional data were collected from 96 participants with ID and 95 of their carers. The service user participants completed an alexithymia questionnaire for children while carers completed the checklist for challenging behaviour and the observer alexithymia scale. Correlational analyses were employed to explore relationships between the variables. RESULTS: The relationship between service user and carer-rated alexithymia was very weak. The analysis did show significant associations between observer-rated alexithymia and challenging behaviour frequency, management difficulty and severity, but there was no significant relationship between challenging behaviour and alexithymia as rated by service users themselves. CONCLUSIONS: This study suggests that observer-rated alexithymia is important in understanding challenging behaviour presented by people with ID. Service user-rated alexithymia had no association with challenging behaviour, in contrast to the results from similar research with other challenging populations.

Retention of corneodesmosomes and increased expression of protease inhibitors in dandruff.

BACKGROUND: Dandruff is a common, relapsing and uncomfortable scalp condition affecting a large proportion of the global population. The appearance of flakes on the scalp and in the hair line, and associated itch are thought to be consequences of a damaged skin barrier, altered corneocyte cohesion and abnormal desquamation in dandruff. The balance between skin proteases and protease inhibitors is essential for driving the key events, including corneodesmosome degradation, in the desquamation process and to maintain stratum corneum (SC) barrier integrity. OBJECTIVES: To investigate the distribution of corneodesmosomes, the key component of the SC cohesivity and barrier function, and the protease inhibitors lympho-epithelial Kazal-type-related inhibitor (LEKTI-1) and squamous cell carcinoma antigen (SCCA1) in the scalp of dandruff-affected participants. METHODS: The methods utilized were immunohistochemistry, scanning immunoelectron microscopy, phase-contrast microscopy, Western blotting and serine protease activity assay on tape-stripped SC or scalp skin biopsies. RESULTS: In SC samples from healthy subjects, corneodesmosomes were peripherally located in the corneocytes. In samples of dandruff lesions, corneodesmosomes were located both peripherally and on the entire surface area of the corneocytes. LEKTI-1 and SCCA1 protein levels and parakeratosis were found to be highly elevated in the lesional samples. CONCLUSIONS: The persistence of nonperipheral corneodesmosomes is a characteristic feature of the perturbed desquamation seen in dandruff. The increased expression levels of LEKTI-1 and SCCA1 are consistent with the view that the dandruff condition is characterized by an imbalance in protease-protease inhibitor interaction in the SC.

Disordered Toll-like receptor 2 responses in the pathogenesis of pulmonary sarcoidosis.

In this study, we hypothesized that the granulomatous disorder sarcoidosis is not caused by a single pathogen, but rather results from abnormal responses of Toll-like receptors (TLRs) to conserved bacterial elements. Unsorted bronchoalveolar lavage (BAL) cells from patients with suspected pulmonary sarcoidosis and healthy non-smoking control subjects were stimulated with representative ligands of TLR-2 (in both TLR-2/1 and TLR-2/6 heterodimers) and TLR-4. Responses were determined by assessing resulting production of tumour necrosis factor (TNF)-α and interleukin (IL)-6. BAL cells from patients in whom sarcoidosis was confirmed displayed increased cytokine responses to the TLR-2/1 ligand 19-kDa lipoprotein of Mycobacterium tuberculosis (LpqH) and decreased responses to the TLR-2/6 agonist fibroblast stimulating ligand-1 (FSL)-1. Subsequently, we evaluated the impact of TLR-2 gene deletion in a recently described murine model of T helper type 1 (Th1)-associated lung disease induced by heat-killed Propionibacterium acnes. As quantified by blinded scoring of lung pathology, P. acnes-induced granulomatous pulmonary inflammation was markedly attenuated in TLR-2(-/-) mice compared to wild-type C57BL/6 animals. The findings support a potential role for disordered TLR-2 responses in the pathogenesis of pulmonary sarcoidosis.

Elevated phenylacetic acid levels do not correlate with adverse events in patients with urea cycle disorders or hepatic encephalopathy and can be predicted based on the plasma PAA to PAGN ratio.

BACKGROUND: Phenylacetic acid (PAA) is the active moiety in sodium phenylbutyrate (NaPBA) and glycerol phenylbutyrate (GPB, HPN-100). Both are approved for treatment of urea cycle disorders (UCDs) - rare genetic disorders characterized by hyperammonemia. PAA is conjugated with glutamine in the liver to form phenylacetyleglutamine (PAGN), which is excreted in urine. PAA plasma levels ≥ 500 µg/dL have been reported to be associated with reversible neurological adverse events (AEs) in cancer patients receiving PAA intravenously. Therefore, we have investigated the relationship between PAA levels and neurological AEs in patients treated with these PAA pro-drugs as well as approaches to identifying patients most likely to experience high PAA levels. METHODS: The relationship between nervous system AEs, PAA levels and the ratio of plasma PAA to PAGN were examined in 4683 blood samples taken serially from: [1] healthy adults [2], UCD patients of ≥ 2 months of age, and [3] patients with cirrhosis and hepatic encephalopathy (HE). The plasma ratio of PAA to PAGN was analyzed with respect to its utility in identifying patients at risk of high PAA values. RESULTS: Only 0.2% (11) of 4683 samples exceeded 500 µg/ml. There was no relationship between neurological AEs and PAA levels in UCD or HE patients, but transient AEs including headache and nausea that correlated with PAA levels were observed in healthy adults. Irrespective of population, a curvilinear relationship was observed between PAA levels and the plasma PAA:PAGN ratio, and a ratio>2.5 (both in µg/mL) in a random blood draw identified patients at risk for PAA levels>500 µg/ml. CONCLUSIONS: The presence of a relationship between PAA levels and reversible AEs in healthy adults but not in UCD or HE patients may reflect intrinsic differences among the populations and/or metabolic adaptation with continued dosing. The plasma PAA:PAGN ratio is a functional measure of the rate of PAA metabolism and represents a useful dosing biomarker.

Adaptive thermoregulation during summer in two populations of an arid-zone passerine.

Heterothermy plays an important role in lowering the costs of thermoregulation in endotherms by reducing water and energy requirements. We tested predictions that birds in arid habitats should express fine-scale variation in their thermoregulatory patterns as a function of prevailing climatic conditions. We assessed effects of air temperature (Tair) and water vapor pressure deficit (D) on body temperature (Tb) in free-living White-browed Sparrow-Weavers (Plocepasser mahali) during summer in two arid habitats in the Kalahari Desert, South Africa, using data from a dry period at a hot, desert site (n=7 birds), and during a dry period (n=4 birds) and a wet period (n=5 birds) at a milder, semi-desert site. The desert birds maintained a significantly higher set-point Tb (41.5 degrees+/-0.2 degrees C, mean-SD) than semi-desert birds (40.2 degrees+/-0.2 degrees C). During the warmest part of day (12:00-18:00 hours), Tb increased significantly during periods of high Tair and/or high humidity, and mean and maximum Tb were up to 1.40 and 2.3 degrees C, respectively, above normal levels. However, as Tair increased, birds at the desert site maintained Tb at or below set-point levels for a greater proportion of the time than birds at the semi-desert site. Birds at the desert site also expressed a greater magnitude of daily heterothermy (heterothermy index, HI=2.4 degrees+/-0.3 degrees C, mean+/-SD) than birds at the semi-desert site: the latter population showed a greater magnitude of heterothermy during a dry period (HI=2.1 degrees+/-0.3 degrees C) than during a wet period (HI=1.6 degrees+/-0.2 degrees C). Birds continued foraging throughout the warmest part of the day, despite the fact that heat dissipation (percentage of time spent panting and wing-spreading) increased significantly with increasing Tair. Our findings reveal that populations can vary in their thermoregulatory responses in both space and time and suggest that small changes in Tair can have significant effects on thermoregulation in free-ranging desert birds, even when Tair

Filaggrin - revisited.

Profilaggrin (proFLG) and its processing products are critical to the health and appearance of skin. The recent identification of loss-of-function filaggrin (FLG) mutations as a predisposing factor in ichthyosis vulgaris and atopic dermatitis has lead to a resurgent interest in this enigmatic protein. Here, we review the literature on the structure and many functions of proFLG, from its role as a filament-aggregating protein and a source of natural moisturizing factor (NMF), to the more recent discoveries of its role in epidermal barrier formation and its more speculative functions as an antimicrobial and sunscreen. Finally, we discuss the relationship of proFLG with dry skin, the influence of moisturizers on NMF generation and speculate on next generation of FLG research.

Urinary phenylacetylglutamine as dosing biomarker for patients with urea cycle disorders.

UNLABELLED: We have analyzed pharmacokinetic data for glycerol phenylbutyrate (also GT4P or HPN-100) and sodium phenylbutyrate with respect to possible dosing biomarkers in patients with urea cycle disorders (UCD). STUDY DESIGN: These analyses are based on over 3000 urine and plasma data points from 54 adult and 11 pediatric UCD patients (ages 6-17) who participated in three clinical studies comparing ammonia control and pharmacokinetics during steady state treatment with glycerol phenylbutyrate or sodium phenylbutyrate. All patients received phenylbutyric acid equivalent doses of glycerol phenylbutyrate or sodium phenylbutyrate in a cross over fashion and underwent 24-hour blood samples and urine sampling for phenylbutyric acid, phenylacetic acid and phenylacetylglutamine. RESULTS: Patients received phenylbutyric acid equivalent doses of glycerol phenylbutyrate ranging from 1.5 to 31.8 g/day and of sodium phenylbutyrate ranging from 1.3 to 31.7 g/day. Plasma metabolite levels varied widely, with average fluctuation indices ranging from 1979% to 5690% for phenylbutyric acid, 843% to 3931% for phenylacetic acid, and 881% to 1434% for phenylacetylglutamine. Mean percent recovery of phenylbutyric acid as urinary phenylacetylglutamine was 66.4 and 69.0 for pediatric patients and 68.7 and 71.4 for adult patients on glycerol phenylbutyrate and sodium phenylbutyrate, respectively. The correlation with dose was strongest for urinary phenylacetylglutamine excretion, either as morning spot urine (r = 0.730, p < 0.001) or as total 24-hour excretion (r = 0.791 p<0.001), followed by plasma phenylacetylglutamine AUC(24-hour), plasma phenylacetic acid AUC(24-hour) and phenylbutyric acid AUC(24-hour). Plasma phenylacetic acid levels in adult and pediatric patients did not show a consistent relationship with either urinary phenylacetylglutamine or ammonia control. CONCLUSION: The findings are collectively consistent with substantial yet variable pre-systemic (1st pass) conversion of phenylbutyric acid to phenylacetic acid and/or phenylacetylglutamine. The variability of blood metabolite levels during the day, their weaker correlation with dose, the need for multiple blood samples to capture trough and peak, and the inconsistency between phenylacetic acid and urinary phenylacetylglutamine as a marker of waste nitrogen scavenging limit the utility of plasma levels for therapeutic monitoring. By contrast, 24-hour urinary phenylacetylglutamine and morning spot urine phenylacetylglutamine correlate strongly with dose and appear to be clinically useful non-invasive biomarkers for compliance and therapeutic monitoring.

Quantifying the effect of urodynamic catheters on urine flow rate measurement.

INTRODUCTION: The effect of urodynamic catheters on urine flow rate (Q(max) ) is well documented but under-researched. Several studies show reduced Q(max) but methodologies and patient demographics differ. The aims of this study were to further quantify the effect of urodynamic catheters on Q(max) and to explore if this was consistent across different urodynamic diagnoses. METHODS: Four groups of 50 consecutive men attending for urodynamic studies (UDS) were retrospectively analyzed: Group 1 comprised 50 men with normal UDS, Group 2 was 50 men with BOO, and Group 3 contained 50 men with detrusor underactivity. Groups 1-3 had UDS performed using both 10 Fr filling and 4 Fr measuring catheters in situ. Group 4 comprised 50 men who had UDS performed with a smaller catheter assembly (8 Fr dual-lumen). Values of Q(max) with and without catheters present were compared using paired Student's t-tests. Differences between groups were compared using ANOVA. RESULTS: Q(max) measured during UDS in men from Groups 1-3 showed a mean reduction of 38% compared to Q(max) from "free" uroflowmetry. ANOVA indicated this reduction was significantly greater among men with normal UDS. Interestingly the group who underwent UDS with a smaller catheter assembly showed no significant reduction in Q(max) measured with catheters in situ. CONCLUSION: Our findings are in line with previous work suggesting that smaller calibre urethral catheters do not cause a significant obstructive effect during voiding. In addition it would appear that the reduction in Q(max) with larger urethral catheters in situ is greatest in those with normal urodynamics.

Measures of the recovery orientation of mental health services: systematic review.

PURPOSE: The review aimed to (1) identify measures that assess the recovery orientation of services; (2) discuss how these measures have conceptualised recovery, and (3) characterise their psychometric properties. METHODS: A systematic review was undertaken using seven sources. The conceptualisation of recovery within each measure was investigated by rating items against a conceptual framework of recovery comprising five recovery processes: connectedness; hope and optimism; identity; meaning and purpose; and empowerment. Psychometric properties of measures were evaluated using quality criteria. RESULTS: Thirteen recovery orientation measures were identified, of which six met eligibility criteria. No measure was a good fit with the conceptual framework. No measure had undergone extensive psychometric testing and none had data on test-retest reliability or sensitivity to change. CONCLUSIONS: Many measures have been developed to assess the recovery orientation of services. Comparisons between the measures were hampered by the different conceptualisations of recovery used and by the lack of uniformity on the level of organisation at which services were assessed. This situation makes it a challenge for services and researchers to make an informed choice on which measure to use. Further work is needed to produce measures with a transparent conceptual underpinning and demonstrated psychometric properties.

Stratum corneum dysfunction in dandruff.

Dandruff is characterized by a flaky, pruritic scalp and affects up to half the world's population post-puberty. The aetiology of dandruff is multifactorial, influenced by Malassezia, sebum production and individual susceptibility. The commensal yeast Malassezia is a strong contributory factor to dandruff formation, but the presence of Malassezia on healthy scalps indicates that Malassezia alone is not a sufficient cause. A healthy stratum corneum (SC) forms a protective barrier to prevent water loss and maintain hydration of the scalp. It also protects against external insults such as microorganisms, including Malassezia, and toxic materials. Severe or chronic barrier damage can impair proper hydration, leading to atypical epidermal proliferation, keratinocyte differentiation and SC maturation, which may underlie some dandruff symptoms. The depleted and disorganized structural lipids of the dandruff SC are consistent with the weakened barrier indicated by elevated transepidermal water loss. Further evidence of a weakened barrier in dandruff includes subclinical inflammation and higher susceptibility to topical irritants. We are proposing that disruption of the SC of the scalp may facilitate dandruff generation, in part by affecting susceptibility to metabolites from Malassezia. Treatment of dandruff with cosmetic products to directly improve SC integrity while providing effective antifungal activity may thus be beneficial.

Assessing changes in stethoscope hygiene during COVID-19: a multi-centre cross-sectional study.

BACKGROUND: The COVID-19 (SARS-CoV-2) pandemic has increased infection control vigilance across several modes of patient contact. However, it is unknown whether hygiene pertaining to stethoscopes, which carry the potential for pathogenic contamination, has also shifted accordingly. AIM: To characterize pandemic-related changes in stethoscope hygiene. METHODS: We surveyed healthcare providers at three major medical centres. Questions quantitatively (Likert scale and frequency) assessed stethoscope hygiene beliefs and practices with two components: before and during COVID-19. Participants were grouped based on performance of optimal stethoscope hygiene (after every patient) before and during COVID-19. Groups were compared using χ2 and analysis of variance (ANOVA). FINDINGS: Of the 515 (10%) who completed the survey, 55 were excluded (N = 460). Optimal hygiene increased from 27.4% to 55.0% (P < 0.001). There were significant increases in Likert scores for all questions pertaining to knowledge of stethoscope contamination (P < 0.001). Belief in stethoscope contamination increased (P < 0.001) despite no change in perceived hygiene education. Resident physicians were less likely compared with attending physicians and nurses to have adopted optimal hygiene during COVID-19 (P < 0.001). CONCLUSION: Despite a positive shift in stethoscope hygiene during COVID-19, optimal hygiene was still only performed by around half of providers. Educational interventions, particularly targeting early-career providers, are encouraged.

Antigen processing and intracellular traffic of antigens and MHC molecules.

Antigen processing leads to binding of antigenic peptides to major histocompatibility complex (MHC) molecules, and these peptide-MHC complexes are recognized by T cells. The class I and class II MHC antigen processing pathways employ different mechanisms and patterns of intracellular transport that allow the two classes to bind and present peptides from different subcellular compartments, determining the source and nature of peptides to be presented.

Processing of Mycobacterium tuberculosis antigen 85B involves intraphagosomal formation of peptide-major histocompatibility complex II complexes and is inhibited by live bacilli that decrease phagosome maturation.

Mycobacterium tuberculosis (MTB) inhibits phagosomal maturation to promote its survival inside macrophages. Control of MTB infection requires CD4 T cell responses and major histocompatibility complex (MHC) class II (MHC-II) processing of MTB antigens (Ags). To investigate phagosomal processing of MTB Ags, phagosomes containing heat-killed (HK) or live MTB were purified from interferon-gamma (IFN-gamma)-activated macrophages by differential centrifugation and Percoll density gradient subcellular fractionation. Flow organellometry and Western blot analysis showed that MTB phagosomes acquired lysosome-associated membrane protein-1 (LAMP-1), MHC-II, and H2-DM. T hybridoma cells were used to detect MTB Ag 85B(241-256)-I-A(b) complexes in isolated phagosomes and other subcellular fractions. These complexes appeared initially (within 20 min) in phagosomes and subsequently (>20 min) on the plasma membrane, but never within late endocytic compartments. Macrophages processed HK MTB more rapidly and efficiently than live MTB; phagosomes containing live MTB expressed fewer Ag 85B(241-256)-I-A(b) complexes than phagosomes containing HK MTB. This is the first study of bacterial Ag processing to directly show that peptide-MHC-II complexes are formed within phagosomes and not after export of bacterial Ags from phagosomes to endocytic Ag processing compartments. Live MTB can alter phagosome maturation and decrease MHC-II Ag processing, providing a mechanism for MTB to evade immune surveillance and enhance its survival within the host.

CpG oligodeoxynucleotides act as adjuvants that switch on T helper 1 (Th1) immunity.

Synthetic oligodeoxynucleotides (ODN) that contain unmethylated CpG motifs (CpG ODN) induce macrophages to secrete IL-12, which induces interferon (IFN)-gamma secretion by natural killer (NK) cells. Since these cytokines can induce T helper 1 (Th1) differentiation, we examined the effects of coadministered CpG ODN on the differentiation of Th responses to hen egg lysozyme (HEL). In both BALB/c (Th2-biased) and B10.D2 (Th1-biased) mice, immunization with HEL in incomplete Freund's adjuvant (IFA) resulted in Th2-dominated immune responses characterized by HEL-specific secretion of IL-5 but not IFN-gamma. In contrast, immunization with IFA-HEL plus CpG ODN switched the immune response to a Th1-dominated cytokine pattern, with high levels of HEL-specific IFN-gamma secretion and decreased HEL-specific IL-5 production. IFA-HEL plus CpG ODN also induced anti-HEL IgG2a (a Th1-associated isotype), which was not induced by IFA-HEL alone. Control non-CpG ODN did not induce IFN-gamma or IgG2a, excepting lesser increases in B10.D2 (Th1-biased) mice. Thus, CpG ODN provide a signal to switch on Th1-dominated responses to coadministered antigen and are potential adjuvants for human vaccines to elicit protective Th1 immunity.

B lymphocytes secrete antigen-presenting vesicles.

Antigen-presenting cells contain a specialized late endocytic compartment, MIIC (major histocompatibility complex [MHC] class II-enriched compartment), that harbors newly synthesized MHC class II molecules in transit to the plasma membrane. MIICs have a limiting membrane enclosing characteristic internal membrane vesicles. Both the limiting membrane and the internal vesicles contain MHC class II. In this study on B lymphoblastoid cells, we demonstrate by immunoelectron microscopy that the limiting membrane of MIICs can fuse directly with the plasma membrane, resulting in release from the cells of internal MHC class II-containing vesicles. These secreted vesicles, named exosomes, were isolated from the cell culture media by differential centrifugation followed by flotation on sucrose density gradients. The overall surface protein composition of exosomes differed significantly from that of the plasma membrane. Exosome-bound MHC class II was in a compact, peptide-bound conformation. Metabolically labeled MHC class II was released into the extracellular medium with relatively slow kinetics, 10 +/- 4% in 24 h, indicating that direct fusion of MIICs with the plasma membrane is not the major pathway by which MHC class II reaches the plasma membrane. Exosomes derived from both human and murine B lymphocytes induced antigen-specific MHC class II-restricted T cell responses. These data suggest a role for exosomes in antigen presentation in vivo.

Phagocytic processing of antigens for presentation by MHC molecules.

Phagocytosis plays a major role in the defence of higher organisms against microbial infection not only by allowing ingested microbes to be destroyed by microbicidal mechanisms, but also by providing the basis for processing of their antigens to forms that generate immune responses. This article examines the role of the phagolysosome in antigen processing, and discusses the contributions of both MHC class II and MHC class I molecules to the presentation of antigens derived from phagocytosed material.

Class II MHC molecules are present in macrophage lysosomes and phagolysosomes that function in the phagocytic processing of Listeria monocytogenes for presentation to T cells.

Phagocytic processing of heat-killed Listeria monocytogenes by peritoneal macrophages resulted in degradation of these bacteria in phagolysosomal compartments and processing of bacterial antigens for presentation to T cells by class II MHC molecules. Within 20 min of uptake by macrophages, Listeria peptide antigens were expressed on surface class II MHC molecules, capable of stimulating Listeria-specific T cells. Within this period, degradation of labeled bacteria to acid-soluble low molecular weight catabolites also commenced. Immunoelectron microscopy was used to evaluate the compartments involved in this processing. Upon uptake of the bacteria, phagosomes containing Listeria fused rapidly with both lysosomes and endosomes. Class II MHC molecules were present in a tubulo-vesicular lysosome compartment, which appeared to fuse with phagosomes, as well as in the resulting phagolysosomes containing internalized Listeria; these compartments were all positive for Lamp 1 and cathepsin D and lacked 46-kD mannose-6-phosphate receptors. In addition, class II MHC and Lamp 1 were co-localized in vesicles of the trans Golgi reticulum, where they were segregated from 46-kD mannose-6-phosphate receptors. Vesicles containing both Listeria-derived components and class II MHC molecules were also observed; some of these may represent vesicles recycling from phagolysosomes, potentially bearing processed immunogenic peptides complexed with class II MHC. These results support a central role for lysosomes and phagolysosomes in the processing of bacterial antigens for presentation to T cells. Tubulo-vesicular lysosomes appear to represent an important convergence of endocytic, phagocytic and biosynthetic pathways, where antigens may be processed to allow binding to class II MHC molecules and recycling to the cell surface.