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A history of colonisation and corresponding traumas has resulted in disparate rates of violence and sexual health inequities among many Native American populations. As a result, Native American adolescents and young adults specifically, experience higher rates of STIs, HIV and unintended pregnancy relative to their non-Hispanic White counterparts. To address these inequities, sexual health education programmes should reflect Native American cultural values and traditional teachings to align with community assets and protective factors. The objective of this study was to describe sexual and reproductive health professionals' perspectives on how trauma collectively affects the sexual health of older adolescents and young adult Native American women between the ages of 15-25 years. We purposively sampled sexual and reproductive health professionals who worked with members of this priority population. Individual in-depth interviews were conducted, recorded, and transcribed. Transcripts were analysed using thematic analysis. The themes identified in the interviews include the impact of trauma and colonisation on sexual health, strategies for combatting trauma, promoting sexual health, and supporting the development of culturally congruent sexual health education curricula. Findings point to the need for culturally relevant, trauma-informed sexual health education interventions to help promote sexual and reproductive health equity for Native American adolescent and young adult women.

The term advanced materials (AM) is used widely to cover a large number of diverse new innovative materials, including nanomaterials, advanced composites, innovative surface coatings, (bio)polymers, porous and particle systems, ceramics, smart and metamaterials and advanced fibres and textiles. With any new materials, there are commercial and performance advantages that need to be balanced with any potential environmental, health and safety issues, for example, around exposure, toxicity, sustainability and waste. Key players in the UK from government bodies, research, measurement and standardisation organisations, academia and industry came together to consider these issues via two online workshops in April 2021 and February 2023. At each event, scene-setting presentations by key experts were followed by discussions addressing salient issues, including, benefits and barriers to AM commercialisation, potential environmental, health and safety issues, and safe(r) by design approaches. The first workshop served as a starting point to share views on the potential societal benefits of AM and perceived obstacles to their wider adoption. The second workshop focused on safety by design, life cycle analysis and challenges faced at different points in the supply chain. In addition to confirming findings from previous studies, these workshops also highlighted specific challenges that are faced by small to medium sized enterprises (SME). These workshops provided a unique opportunity for policy makers, regulators, standardisation bodies, funding bodies and academia to understand the concerns of industry and researchers, who develop and work with AM. This included what they felt would help support them in their aims of developing innovative, commercially successful, safe and sustainable AM., Competing Interests: There are no conflicts of interest to declare., (This journal is © The Royal Society of Chemistry.)

This review is about the ethical use of sensors to monitor occupational exposure to hazards. It considers whether the same or different, ethical measures apply to using sensors, compared to conventional hazard monitoring surveys. To undertake the review, subject experts developed a research question, identified suitable search terms, and set the scope of these searches. Candidate research papers dating from 2000 to mid-2022 that met inclusion criteria were identified and reviewed by each author. Ethical concerns were identified by the authors of studies in which sensors were used to monitor employee health and well-being, but most of the studies that used them to monitor employee exposure to hazards focused on the technical aspects of their deployment. These ethical concerns included questions about employee rights and privacy, the anonymity of the data collected with sensors, and how the security of this data is managed. The review considers ethical standards and codes of practice for occupational hygiene work and the ethical risks when sensors are used to gather data. Sensors may provide insight into occupational exposure to hazards, but their use is not always adequately explained to employees by those managing this monitoring work. The ethical concerns identified were relevant to many areas of industrial hygiene work, but more studies are required that consider the ethical use of sensors in workplaces. Studies that monitored employee health, well-being, and productivity, identified ethical risks in using sensors to monitor these endpoints. An ethical framework and checklist for hygienists are proposed including a set of questions that consider the risks of using sensors to monitor occupational hazards. Industrial hygiene professional bodies provide ethical codes of practice for their members but may also need to consider the implications of using sensors in workplaces. Ethical standards support the collection of industrial hygiene exposure data whilst maintaining the privacy rights of employees.

Background: Although the literature suggests that medication-assisted treatment (MAT) is an effective treatment for opioid use disorder, limited studies have assessed the prevalence or the association between MAT use and sexual identity, mental health, or substance use disorder among a nationally representative sample. We assessed the prevalence and association of opioid MAT use between sexual identity, depressive disorder symptoms, alcohol use dependence, and marijuana use dependence in the United States., Methods: We used the 2019 National Survey on Drug Use and Health public-use data on adults aged 18-64 years ( N = 38,841) to conduct a weighted multivariable logistic regression analysis., Results: A total of 4.80% and 2.32% of the population identified as bisexual and lesbian/gay, respectively. About 0.31% (612,750 people) of the population reported receiving opioid MAT, 3.73% had alcohol use dependence, 1.42% had marijuana use dependence, and 9.13% had major depressive episode (MDE) symptoms. Of those who had received opioid MAT, 0.57% were bisexuals and 1.07% were lesbians/gays, 0.65% were people with alcohol use dependence, 2.32% with marijuana use dependence, and 1.59% with MDE symptoms. Lesbian/gay individuals were more likely to receive opioid MAT (AOR = 3.43, 95% CI = 1.42, 8.25) compared to heterosexual individuals. The odds were higher for people with marijuana use dependence (AOR = 3.44, 95% CI = 1.47, 8.06) and MDE symptoms (AOR = 5.22, 95% CI = 3.46, 7.89) than their counterparts., Conclusions: In this study, sexual minorities, people with MDE symptoms, and those dependent on marijuana use were more likely to receive opioid MAT, suggesting the need to investigate further opioid use disorder symptoms and their risk factors among these populations., Competing Interests: Competing interests The authors have no competing interests to declare that are relevant to the content of this article.

Background: American Indian and Alaska Native (AI/AN) youth experience serious disparities in sexual and reproductive health, including the highest teen birth rate among racial/ethnic groups, and disproportionate rates of sexually transmitted infections (STI), including HIV. A growing number of evidence-based programs (EBPs) that integrate the strengths and cultural teachings of Native communities exist. Yet, multiple factors, including lack of trained personnel, limited resources, and geographic isolation, may hinder their adoption and implementation. Innovative implementation strategies that facilitate the adoption and implementation of sexual health EBPs in Native communities may help reduce these disparities., Methods: We applied Implementation Mapping, a systematic planning framework that utilizes theory, empirical evidence, and community input, to adapt a theory-based, online decision support system, iCHAMPSS (CHoosing And Maintaining Effective Programs for Sex Education in Schools), to support underlying dissemination and implementation processes unique to Native communities. We used an iterative design process, incorporating input from Native practitioners and academicians, to ensure that the adapted decision support system reflects cultural identification, community values, and experiences., Results: Grounded in diffusion of innovations, organizational stage theory, and social cognitive theory, the Healthy Native Youth Implementation Toolbox supports Native practitioners through five phases (Gather, Choose, Prepare, Implement, and Grow) to adopt, implement, and maintain a culturally-relevant, age-appropriate sexual health EBP. The Toolbox provides tools, ready-to-use templates, and guidance to plan, implement, and grow a culturally-relevant adolescent health program with their Tribe or community. Hosted within the Healthy Native Youth website (www.healthynativeyouth.org), the Toolbox comprises: (1) a curriculum portal with access to 15 culturally-relevant, age-appropriate evidence-based health promotion programs for AI/AN youth; (2) a "resource library" comprising 20+ support tools, templates, and links to external resources, and (3) "stories from the field" comprising testimonials from experienced Native educators, who have implemented sexual health programs., Conclusion: There is a continued need to design, test, and evaluate D&I strategies that are relevant to Native communities. The Healthy Native Youth Implementation Toolbox contributes to the dissemination and implementation of evidence-based, culturally-relevant sexual health education programs in diverse Native communities. Implementation Mapping provided a systematic approach to guide the adaptation process and integrate community voice with the ultimate goal of enhancing sexual health equity among AI/AN youth., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationship that could be construed as a potential conflict of interest., (Copyright © 2022 Markham, Rushing, Manthei, Singer, Jessen, Gorman, Peskin, Hernandez, Sacca, Evans, Luna-Meza, Merritt and Shegog.)

Dermal exposure to metal allergens can lead to irritant and allergic contact dermatitis (ACD). In this paper we present a mathematical model of the absorption of metal ions, hexavalent chromium and nickel, into the viable epidermis and compare the localised irritant and T-lymphocyte (T-cell) mediated immune responses. The model accounts for the spatial-temporal variation of skin health, extra and intracellular allergen concentrations, innate immune cells, T-cells, cytokine signalling and lymph node activity up to about 6 days after contact with these metals; repair processes associated with withdrawal of exposure to both metals is not considered in the current model, being assumed secondary during the initial phases of exposure. Simulations of the resulting system of PDEs are studied in one-dimension, i.e. across skin depth, and three-dimensional scenarios with the aim of comparing the responses to the two ions in the cases of first contact (no T-cells initially present) and second contact (T-cells initially present). The results show that on continuous contact, chromium ions elicit stronger skin inflammation, but for nickel, subsequent re-exposure stimulates stronger responses due to an accumulation of cytotoxic T-cell mediated responses which characterise ACD. Furthermore, the surface area of contact to these metals has little effect on the speed of response, whilst sensitivity is predicted to increase with the thickness of skin. The modelling approach is generic and should be applicable to describe contact dermatitis from a wide range of allergens.

The KG1 myeloid leukaemia was used as source of dendritic cells (DC) to discriminate between respiratory and contact sensitising chemicals. A cocktail of cytokines was used to differentiate KG1 to dendritic like cells (termed dKG1) and the effects of nine chemicals (respiratory and contact sensitisers) and an irritant control on surface marker expression, 'antigen presenting' function and cytokine expression investigated. The stability of these chemicals when dissolved was characterised using MALDI ToF MS. A Hill plot model was used with the cellular viability data to quantify the lethal dose 50% (LD50) and a maximum sub toxic concentration of each chemical defined. Cytokine expression by the treated dKG1 was quantified using multiplex immunobead analysis. Whilst dKG1 cells were morphologically similar to DCs, expression of specific surface markers was not typical for DCs derived from healthy precursor cells. When the chemicals were applied at defined sub toxic doses no effects on dKG1 phenotype, function, or cytokine expression, attributable to the sensitisation properties were discriminated. However, dKG1 cells were much more sensitive to the toxic effects of these chemicals compared to the parent KG1 cells. Only 4 of the 9 chemicals tested were stable when dissolved indicating that the effect of sensitising chemicals on antigen presenting cells may be related to species other than the parent compound., (Crown Copyright © 2015. Published by Elsevier Ireland Ltd. All rights reserved.)

Background: Since the mid-1990s, outbreaks of asthma and extrinsic allergic alveolitis (EAA) have been identified in workers exposed to metalworking fluids (MWFs). The cause of these outbreaks remains to be determined., Aims: To identify and review all previously published occupational lung disease case definitions and diagnostic criteria that have been utilized during MWF outbreak investigations., Methods: Respiratory outbreaks due to MWFs were identified by a systematic literature search for articles published between 1990 and October 2011. Investigations reporting the usage of disease case definitions or diagnostic criteria for respiratory disease were reviewed and summarized., Results: The literature search identified 35 papers relating to 27 outbreaks of respiratory disease in MWF-exposed workers. Fourteen case definitions for MWF-related respiratory disease were identified: seven for EAA, five for occupational asthma and one each for humidifier fever and industrial bronchitis. A single paper was identified where any comparison of different disease case definitions (for EAA) had been performed., Conclusions: A range of case definitions and diagnostic criteria for MWF respiratory disease have been utilized in outbreak investigations, but the majority have been produced for individual outbreak investigations without previous validation. It may be difficult to compare the findings of future workplace studies without a more standardized approach to case identification and diagnosis.

A sound knowledge base is required to target resources to reduce workplace exposure to carcinogens. This project aimed to provide an objective estimate of the burden of cancer in Britain due to occupation. This volume presents extensive analyses for all carcinogens and occupational circumstances defined as definite or probable human occupational carcinogens by the International Agency for Research on Cancer. This article outlines the structure of the supplement - two methodological papers (statistical approach and exposure assessment), eight papers presenting the cancer-specific results grouped by broad anatomical site, a paper giving industry sector results and one discussing work-related cancer-prevention strategies. A brief summary of the methods and an overview of the updated overall results are given in this introductory paper. A general discussion of the overall strengths and limitations of the study is also presented. Overall, 8010 (5.3%) total cancer deaths in Britain and 13,598 cancer registrations were attributable to occupation in 2005 and 2004, respectively. The importance of cancer sites such as mesothelioma, sinonasal, lung, nasopharynx, breast, non-melanoma skin cancer, bladder, oesophagus, soft tissue sarcoma and stomach cancers are highlighted, as are carcinogens such as asbestos, mineral oils, solar radiation, silica, diesel engine exhaust, coal tars and pitches, dioxins, environmental tobacco smoke, radon, tetrachloroethylene, arsenic and strong inorganic mists, as well as occupational circumstances such as shift work and occupation as a painter or welder. The methods developed for this project are being adapted by other countries and extended to include social and economic impact evaluation.

Introduction: Potential demographic risk factors for outbreaks of respiratory disease due to water-based metalworking fluids (MWFs) were investigated through systematic review of published outbreak investigations., Methods: Search terms were selected by a multidisciplinary team, assisted by an experienced library information service. Several computerized literature databases were searched for articles published between January 1990 and October 2011, relating to ill health outbreaks due to MWFs. Papers meeting the search criteria were reviewed in detail, and their references checked for additional articles. Study design and demographic details of the outbreak were extracted from the selected articles and entered into standardized evidence tables., Results: Thirty-five articles relating to investigations of 27 outbreaks of respiratory ill health attributed to MWF exposure were identified. The majority of reports were case series of disease or observational cross-sectional studies of symptoms and hygiene measurements. Eight of the outbreak investigations included an element of case-control analysis. Most outbreaks were from the USA, had occurred in large car- or aeronautical-manufacturing plants, and were associated with the use of central shared sumps. Hygiene studies have not demonstrated consistent risk factors for respiratory outbreaks, in terms of the type of MWF utilized, degree of microbial contamination, or levels of personal exposure. Six studies were identified that found workers with MWF exposure during outbreaks were more likely to report respiratory or systemic symptoms than unexposed control workers. Six case-control analyses were also identified that found workers with extrinsic allergic alveolitis (EAA) were more likely to demonstrate certain immune responses to microbial contaminants and/or used MWFs than workers without EAA., Conclusion: Despite a number of detailed workplace and immunological studies of asthma and alveolitis outbreaks in MWF-exposed workforces, our understanding of their aetiology remains limited.

A mathematical model describing the main mechanistic processes involved in keratinocyte response to chromium and nickel has been developed and compared to experimental in vitro data. Accounting for the interactions between the metal ions and the keratinocytes, the law of mass action was used to generate ordinary differential equations which predict the time evolution and ion concentration dependency of keratinocyte viability, the amount of metal associated with the keratinocytes and the release of cytokines by the keratinocytes. Good agreement between model predictions and existing experimental data of these endpoints was observed, supporting the use of this model to explore physiochemical parameters that influence the toxicological response of keratinocytes to these two metals.

The European Centre for Ecotoxicology and Toxicology of Chemicals (ECETOC) convened a workshop in June 2005 to address the speculation that exposure to specific chemicals, and/or chemical pollutants in general, may play an important role in the increased prevalence of allergy and asthma in 'westernized' societies. This paper summarises one perspective arrived at during this workshop. It was acknowledged that certain chemicals and certain types of pollution might trigger or exacerbate asthmatic reactions in sensitised subjects. However, overall levels of pollution appear not to have had a major impact upon the prevalence of atopic allergy. Epidemiological studies suggest that pollution may in some circumstances protect from acquisition of sensitisation. Increasing exposure to household chemicals may enhance pre-existing allergies, but evidence for their causation of allergy is lacking. Other risk factors considered included societal dietary changes and exposure to endotoxins. Future research needs were identified which included epidemiological studies employing exposure and biomonitoring data, studies on domestic exposure to chemicals and their association with the incidence of allergy and asthma, and prospective birth cohort studies employing well-defined aspects of lifestyle, diet, chemical and endotoxin exposure as factors that may drive susceptibility to allergy and asthma., ((c) 2007 John Wiley & Sons, Ltd.)

Background: Exposure to metal working fluid (MWF) has been associated with outbreaks of extrinsic allergic alveolitis (EAA) in the USA, with bacterial contamination of MWF being a possible cause, but is uncommon in the UK. Twelve workers developed EAA in a car engine manufacturing plant in the UK, presenting clinically between December 2003 and May 2004. This paper reports the subsequent epidemiological investigation of the whole workforce. The study had three aims: (1) to measure the extent of the outbreak by identifying other workers who may have developed EAA or other work-related respiratory diseases; (2) to provide case detection so that those affected could be treated; and (3) to provide epidemiological data to identify the cause of the outbreak., Methods: The outbreak was investigated in a three-phase cross-sectional survey of the workforce. In phase I a respiratory screening questionnaire was completed by 808/836 workers (96.7%) in May 2004. In phase II 481 employees with at least one respiratory symptom on screening and 50 asymptomatic controls were invited for investigation at the factory in June 2004. This included a questionnaire, spirometry and clinical opinion. 454/481 (94.4%) responded and 48/50 (96%) controls. Workers were identified who needed further investigation and serial measurements of peak expiratory flow (PEF). In phase III 162 employees were seen at the Birmingham Occupational Lung Disease clinic. 198 employees returned PEF records, including 141 of the 162 who attended for clinical investigation. Case definitions for diagnoses were agreed., Results: 87 workers (10.4% of the workforce) met case definitions for occupational lung disease, comprising EAA (n = 19), occupational asthma (n = 74) and humidifier fever (n = 7). 12 workers had more than one diagnosis. The peak onset of work-related breathlessness was Spring 2003. The proportion of workers affected was higher for those using MWF from a large sump (27.3%) than for those working all over the manufacturing area (7.9%) (OR = 4.39, p<0.001). Two workers had positive specific provocation tests to the used but not the unused MWF solution., Conclusions: Extensive investigation of the outbreak of EAA detected a large number of affected workers, not only with EAA but also occupational asthma. This is the largest reported outbreak in Europe. Mist from used MWF is the likely cause. In workplaces using MWF there is a need to carry out risk assessments, to monitor and maintain fluid quality, to control mist and to carry out respiratory health surveillance.

This study was carried out to assess the effects of chromium and nickel upon isolated keratinocytes as an in vitro model of human skin. Keratinocytes were isolated from healthy volunteer skin samples of unknown metal sensitivity (n=10) and were compared with cells from patient biopsies of known metal sensitivity (n=7). Cells were dosed with a concentration range of nickel and chromium (0-10,000 microM) and cellular mitochondrial activity, viability, metal uptake and cytokine release were measured. Responses of primary versus passaged keratinocytes were also compared. Toxicity data from primary and passaged keratinocytes was statistically analysed by the non-linear Hill Plot model. Results showed that hexavalent chromium was significantly more cytotoxic, associated more with keratinocytes and induced a dose dependant release of IL-1alpha compared to nickel. Significant differences were observed between primary and passaged keratinocytes with regard to the toxicity of chromium and nickel and variation of response. No differences were observed in the cytotoxicity or cytokine release induced by chromium or nickel for the known sensitised biopsy patient samples (n=4) compared to patch test negative controls (n=3). The results from this study suggest human keratinocytes in vitro respond very differently to chromium and nickel.

Objectives: The aim of this study was to investigate whether prelabour ruptured membranes at term display increased proteolytic activity and to determine whether regional structural alterations within membranes are reflective of regional variations in proteolytic activity., Study Design: Multiple amnion samples were collected from 37 women with prelabour membrane rupture and 37 women whose membranes ruptured spontaneously during labour. In all cases the gestation was greater than 37 weeks. Substrate zymography was used to qualitatively assess gelatinase and serine protease involvement. General protease activity (metallo, serine, acid and sulfhydryl) was measured quantitatively by fluorescent substrate cleavage., Results: Substrate zymography revealed no active gelatinases or serine proteases. Fluorescent studies of general protease activity showed no significant difference between the groups and no significant regional variation., Conclusions: Gelatinase and serine protease activity do not play a major role in the formation of a membrane rupture initiation site or in prelabour membrane rupture at term.

Objectives: Enzymes are commonly used in the baking industry, as they can improve dough quality and texture and lengthen the shelf life of the final product. There is little published information highlighting exposure to enzymes (other than fungal alpha-amylase) in the baking industry, therefore the purpose of this study was to identify antibodies and develop assays for the measurement of a variety of such enzymes in samples of airborne flour dust., Methods: Polyclonal antibodies to bacterial amylase, glucose oxidase and amyloglucosidase were identified and developed into ELISA assays. The assays showed limited cross-reactivity with other enzymes commonly used in the baking industry., Results: We measured levels of airborne enzymes in 195 personal air samples taken from a sample of 55 craft baking establishments. We were able to detect amyloglucosidase in 9% (16/184) of the samples, fungal alpha-amylase in 6% (11/171), bacterial alpha-amylase in 7% (13/195). However, we were unable to detect glucose oxidase in any of the samples. Measurements for protease enzymes were not carried out. Median levels in detectable samples of amyloglucosidase, fungal alpha-amylase and bacterial amylase were similar at 10.3, 5.3 and 5.9 ng/m(3), respectively. These figures represent the total enzyme protein (active and inactive) measured., Conclusions: There are few data in the literature regarding sensitization and exposure-response relationships to these enzymes, and indeed there is often a lack of information within the industry as to the precise enzyme content of particular baking ingredients. As a precautionary measure, all enzymes are regarded as having the potential to cause respiratory sensitization. Consequently, exposures need to be controlled to as low a level as reasonably practicable, and future investigation may highlight the importance of measuring a variety of enzyme exposures and standardizing these methodologies to inform approaches to adequate control.

RSV causes annual epidemics of bronchiolitis in winter months resulting in the hospitalization of many infants and the elderly. Dendritic cells (DCs) play a pivotal role in coordinating immune responses to infection and some viruses skew, or subvert, the immune functions of DCs. RSV infection of DCs could alter their function and this could explain why protection after natural RSV infection is incomplete and of short duration. In this study, this interaction between DCs and RSV was investigated using a human primary culture model. DCs were generated from purified healthy adult volunteer peripheral blood monocytes. Effects of RSV upon DC phenotype with RSV primed DCs was measured using flow cytometry. Changes to viability and proliferation of cocultured DCs and T-cells were determined using microscopy with fluorescent dyes (Hoechst 33342 and propidium iodide). DC maturation was not prevented by the RSV challenge. RSV infected a fraction of DCs (10-30%) but the virus replicated slowly in these cells with only small reduction to cell viability. DCs challenged with RSV stimulated T-cell proliferation less well than lipopolysaccharide. This is the first study to demonstrate RSV infection of human monocyte derived DCs and suggests that the virus does not significantly interfere with the function of these cells and potentially may promote cellular rather than humoral responses.

Respiratory syncytial virus infects almost all children by 2 years of age. Neutrophils are the predominant airway leucocytes in RSV bronchiolitis and they are activated in the presence of infection. However it is not clear whether RSV can directly signal to activate neutrophil cytotoxic function. To investigate this we have used a preparation of RSV washed using a new centrifugal diafiltration method to rapidly remove inflammatory molecules produced by the epithelial cells used to propagate the RSV stock. Human neutrophils were isolated from peripheral blood and activated with either the unwashed crude RSV preparations or the purified intact RSV. Neutrophils were also challenged with purified RSV G-glycoprotein. The effect of challenging human neutrophils with these preparations of intact RSV, or the RSV G-glycoprotein, was assessed by measuring the cell surface expression of CD11b and CD18b, the phagocytic oxidative burst, and intracellular release of calcium pools. Neutrophils challenged with the washed RSV exhibited significantly lower activation of surface marker expression (P < 0.001) and oxidative burst (P < 0.001) than those challenged with unwashed virus or with virus free supernatant. There was no increase in intracellular calcium release on exposure to the washed RSV. Purified G glycoprotein did not stimulate neutrophils, whilst the use of a blocking antibody to the F protein did not prevent unwashed RSV from activating cytotoxic responses. These results suggest that neutrophils have no innate signalling system that recognizes RSV but they are activated at sites of RSV infection as a result of the cytokines and inflammatory molecules released by virally infected cells.

Prelabor rupture of the fetal membranes affects approximately 10% of women at term, resulting in an increased risk of maternal and neonatal infection. Evidence suggests that membrane rupture is related to biochemical processes involving the extracellular matrix of the membranes. We tested the hypothesis that prelabor ruptured membranes are characterized by reduced collagen concentrations, altered collagen cross-link profiles, and increased concentrations of biomarkers of oxidative damage. We also set out to determine whether these effects are modulated by ascorbic acid status. In a case-control study, we explored the role that ascorbic acid, oxidative stress, collagen, and collagen cross-links play in determining membrane integrity and developed a functional assay to assess membrane proteolytic susceptibility. Prelabor ruptured membrane had a reduced ascorbic acid concentration in comparison with controls while protein carbonyl and malondialdehyde concentrations were increased. Collagen concentrations were also reduced in prelabor ruptured membrane, and while the concentration of collagen cross-links was not significantly different between prelabor and timely ruptured membrane, there was a regional variation in cross-link ratio within the amniotic sac. Proteolytic resistance in vitro was reduced in prelabor ruptured membrane and also exhibited regional variation within the amniotic sac. Our findings are strongly supportive of a role for the enhanced degradation of membrane collagen in the determination of prelabor rupture of fetal membranes. The formation of the rupture initiation site is a function of a regional variation in collagen cross-link ratio. Tissue ascorbic acid status may be an important mediator of these processes.

Inflammatory mediators have been reported to promote malignant cell growth, invasion and metastatic potential. More specifically, we have recently reported that tumour necrosis factor alpha (TNF-alpha) increases melanoma cell attachment to extracellular matrix (ECM) substrates and invasion through fibronectin. In this study, we extend these investigations asking specifically whether the TNF-alpha effect on cell invasion and migration involves activation of proteolytic enzymes. We examined the effect of TNF-alpha on melanoma expression/activation of type IV gelatinases matrix metalloproteinases 2 and 9 (MMPs -2 and -9) and general proteolytic enzymes. Stimulation with TNF-alpha significantly increased both melanoma cell migration at 24 h (+21%) and invasion through fibronectin (+35%) but did not upregulate/activate the expression of latent MMP-2 constitutively produced by these cells and did not upregulate their general protease activity. However, the increased cell migration and invasion through fibronectin observed following stimulation with TNF-alpha were inhibited by the general protease inhibitor alpha(2) macroglobulin. These findings suggest that the promigratory and proinvasive effect of TNF-alpha on this melanoma cell line may be mediated to some extent by induction of localised cell membrane-bound degradative enzyme activity, which is not readily detected in biochemical assays.

An increase in the size and cellularity of duodenal crypts and a decreased incidence of bifurcating crypts is observed in response to very short-term feeding of a riboflavin-deficient diet to weanling rats. A study was conducted to determine whether the absence of riboflavin in the lumen of the small intestine impairs gastrointestinal development. Forty-eight female weanling Wistar rats were allocated to one of two treatment regimens, to receive either a riboflavin-deficient diet and a daily intraperitoneal injection of flavin mononucleotide (luminally deficient group) or a complete diet and a daily intraperitoneal injection of saline (control group). Animals were killed at 93, 141, or 165 hr from feeding. The flavin injection regimen maintained normal systemic riboflavin status in the luminally deficient group. In this group, however, crypt hypertrophy and reduced crypt bifurcation were evident by 141 hr of luminal riboflavin deprivation. The absence of riboflavin in the duodenal lumen impairs normal development, suggesting that a crypt sensing mechanism may be involved in the response to riboflavin deficiency.

Aim: To assess the potential of gingival crevicular fluid (GCF) from adult periodontitis patients and Porphyromonas gingivalis proteases to activate matrix metalloproteinase 2 (MMP-2) in vitro., Material and Methods: GCF samples were collected from each of 15 adult periodontitis patients, from a clinically healthy site, a deep (>6 mm) bleeding site, and a deep nonbleeding site. The GCF samples were examined for general proteolytic activity, gelatinolytic activity and ability to activate pro-MMP-2 by zymography. Ultrasonic extracts of a range of clinical isolates of P. gingivalis cells and purified arg- and lys-gingipains were also assessed for their ability to activate pro-MMP-2., Results: GCF from deep nonbleeding sites showed higher general proteolytic activity than samples from deep bleeding and healthy sites but this did not reach statistical significance. Pefabloc, a general serine protease inhibitor, inhibited the majority (92%) of the proteolytic activity. GCF samples contained neutrophil MMP-9 in its latent form in 93% of the samples, and in its activated form in 40% of the samples. In contrast, MMP-2 was present in only trace amounts in 9% of the samples. When latent MMP-2 was added to these GCF samples, it was converted to the activated form (59 kDa) in 68% of the samples. Lower molecular weight (55 and 45 kDa) activated forms also appeared in 53% of the samples, particularly those from deep sites. Activation to the 55 and 45 kDa forms was inhibited by MSAAPket (a neutrophil elastase inhibitor), whereas Pefabloc completely inhibited the activation of latent MMP-2. All ultrasonic extracts of P. gingivalis activated latent MMP-2 in a concentration- and time-dependent manner. Also, latent MMP-2 was activated by purified arg-gingipain but less efficiently by lys-gingipain., Conclusion: These findings suggest that P. gingivalis arg-gingipain and neutrophil elastase present in GCF can activate latent MMP-2, which may contribute in vivo to local periodontal tissue destruction.

Large numbers of neutrophils in the airway of infants infected by respiratory syncytial virus (RSV) are recruited by chemokines, such as interleukin-8, and specific inflammatory molecules can delay apoptosis increasing their longevity. The aim of this study was to investigate whether airway secretions in RSV bronchiolitis contain factors that influence neutrophil apoptosis. Nasal lavage fluid (NLF) was obtained from 24 infants with RSV bronchiolitis (31 infant controls and 12 adults). Neutrophils isolated from healthy adult volunteers were incubated with the NLF in Dulbecco modified Eagle medium (DMEM) for 24 h, and apoptosis and necrosis were quantified using Hoechst 33342 and propidium iodide viability dyes. The presence of putative factors that delay neutrophil apoptosis was investigated using inhibitors to leukotriene-B4, lipopolysaccharide and the IL-8 receptor CXCR2, and blocking antibodies to granulocyte-monocyte colony-stimulating factor. Characterisation of NLF involved tests of thermal instability, proteolysis, deoxyribonuclease digestion and molecular filtration. NLF from infants with RSV bronchiolitis and controls significantly delayed neutrophil apoptosis, whereas NLF from healthy adults did not. None of these inhibitor molecules blocked this delay in apoptosis but activity was heat liable and >3 kDa. The study showed that nasal lavage fluid from infants significantly delays neutrophil apoptosis. The speculation is that the prolonged survival of neutrophils in the infant airway contributes to the characteristic accumulation of neutrophils in the airways of infants with respiratory infections.

Nonantimicrobial effects of antibiotics may contribute to their activity in the treatment of infective airway disease. The aim of this study was to identify antibiotics used for the treatment of infection in cystic fibrosis that may alter the activity of human neutrophil elastase (HNE) and Pseudomonas aeruginosa elastase (PE). The effect of antibiotics on the activity of purified HNE and PE, and HNE in sputum was assessed using colourimetric and fluorescent substrate assays by kinetic measurements, and by examining the interaction of HNE with inhibitors. Ceftazidime, tobramycin, and gentamycin slightly inhibited purified HNE activity whereas erythromycin and colistin significantly stimulated purified HNE and PE (395 and 557%, respectively). However, only colistin increased HNE activity in sputum (+102%) and was therefore studied in more detail. This increase in activity was not due an interference with the specific inhibition of HNE by alpha1-antitrypsin but colistin was found to reverse the inhibitory effects of small molecular weight molecules like heparin. Colistin increases the activity of human neutrophil elastase and Pseudomonas aeruginosa elastase, two proteases that contribute to the pathogenesis of cystic fibrosis airway disease.

The aim of this present study was to identify the earliest point at which riboflavin deficiency affects post-weaning bowel development in rats. After weaning, eighty Wistar rats were weight-matched as pairs, one animal being fed a normal synthetic diet and the other being fed the same diet but deficient in riboflavin. Body weight, feeding and rates of growth were monitored and eight pairs of animals were taken for analysis at 45, 69, 93, 117 and 141 h. Riboflavin status was monitored by determining the erythrocyte glutathione reductase activation coefficient (EGRAC), and hepatic flavins were measured by a fluorescence assay. Changes to the number and dimensions of villi and crypts in the duodenum were determined, as well as crypt division (bifurcation) and the DNA synthesis index of the crypt epithelium by bromodeoxyuridine (BrdU) labelling. Riboflavin deficiency was established in the experimental rats, as demonstrated by a significant increase in EGRAC after 45 h (P<0.001) and decreased liver flavins after 96 h (P<0.001). After 96 h a significant increase in the size and cellularity of the crypts (P<0.001 in both cases) was seen in these riboflavin-deficient animals, with a decreased incidence of bifurcating crypts and of BrdU-labelled cells. No changes to villus number or size were observed. The present study has demonstrated that developmental changes to the duodenal crypt arise shortly after circulating riboflavin measurements show evidence of deficiency. These changes primarily affect cell proliferation and crypt bifurcation, and precede long-term changes such as the reduction of villus number.

Aims: To assess the implications of detection of interleukin 8 (IL-8) in urine., Methods: IL-8 was measured by immunoassay in all 305 urine samples from children aged 0-18.4 years received by our microbiology laboratory during four weeks, with a retrospective structured case note audit for all those in whom IL-8, white cells, or bacteria were detected. Patients were divided into three groups: urinary tract infection (UTI), at least one sample with >/=5 leucocytes x 10(9)/l and >/=10(5) cultured bacteria/ml; possible UTI, at least one sample with >/=5 leucocytes x 10(9)/l or >/=10(5) cultured bacteria/ml but not both; UTI unlikely, sample(s) with <5 leucocytes x 10(9)/l and <10(5) cultured bacteria/ml. Medical records were sought for all in groups 1 (14/14 found) and 2 (18/21 found) and those in group 3 (41/59 found) in whose urine any leucocytes, cultured bacteria, or IL-8 were detected., Results: IL-8 was detected in 58/305 samples from 48/264 patients. IL-8 was detected in at least one urine sample from 13/14 patients with confirmed UTI (group 1); in 11/21 patients with possible UTI (group 2), of whom two were treated as UTI; and in 23/228 patients without UTI. Using a cut off of 200 pg/ml, urine IL-8 had a sensitivity of 93% and a specificity of 90% for diagnosing UTI., Conclusions: Urine IL-8 is a sensitive test for UTI, but is poorly specific as it is also present in a variety of other infectious and inflammatory disorders.

Adherence of Escherichia coli to urinary tract epithelium induces neutrophil migration across the uroepithelium to combat bacterial infection. Neutrophil adherence to the apical membrane of uroepithelial cells may be an important factor for bacterial clearance. We used an in vitro model of urinary tract infection to examine the effects of uropathogenic E. coli on neutrophil adhesion to the uroepithelial cell line RT4. We found that distinct clinical isolates caused different levels of neutrophil adherence. One particular isolate caused significant neutrophil adhesion in a dose- and time-dependent manner. The neutrophil adhesion-promoting effect induced by this isolate was not caused by bacterial secreted products, suggesting that contact between intact E. coli and uroepithelial cells is required for promoting neutrophil adhesion. This adhesion was almost exclusively mediated by CD11b/CD18, suggesting that E. coli upregulates CD11b/CD18 counterligands on the uroepithelial surface. These data suggest that certain uropathogenic E. coli selectively promote adhesion of neutrophils to ligands on uroepithelial cells by a CD11b/CD18-dependent mechanism.

Rat intestinal fibroblast lines (F1:G9 and A1:F1) differing in their potential to support intestinal mucosal development were marked with reporter genes to investigate their differentiation potential. The fibroblasts were transfected with plasmids expressing either beta-galactosidase (with or without a nuclear localisation signal) or green fluorescent protein (GFP). Transfection using Tfx50 or Fugene was more efficient than electroporation. The expression of beta-galactosidase was more stable and stronger than GFP. Cells were optimally labelled using the plasmid pL27B-GAL, and sub-clones with a strong and uniform nuclear expression of beta-galactosidase were isolated. These clones expressed beta-galactosidase even after prolonged passage in the absence of selection. The beta-galactosidase tagged lines (F1:G9gal and A1:F1gal) retained the morphological characteristics, viability and differentiation properties of the parental non-transfected lines. In co-culture with a colorectal tumour cell line Caco-2, the F1:G9gal and A1:F1gal cells differed in their morphological organisation but this did not change their expression of smooth muscle alpha-actin., (Copyright 2001 Academic Press.)

In Wilson's disease and Indian childhood cirrhosis (ICC) copper accumulates in the liver resulting in poor hepatocyte regeneration and fibrosis. An inhibition of hepatocyte proliferation and an increase in cell death could account for these outcomes. To establish how the toxicity of this metal ion impacts upon the proliferation and viability of the HepG2 cells they were cultured in 4-32 microM copper(II) sulphate (CuSO4)). These levels were comparable to the circulatory and tissue concentrations of copper recorded for these two diseases. Specific uptake comparable to levels of copper recorded in the livers of patients with Wilson's disease and ICC was measured in the HepG2 cells. After 48 h acid vesicle function increased from 4 to 32 microM Cu2+ but significantly declined at 64 microM compared to the controls. Lysosomal acid phosphatase showed a concentration dependent decline in activity at 72 h. Cellls exposed to 64 microM Cu2+ had a potential doubling time (Tpot) 21 h longer than the control cells due to a prolonged DNA synthesis phase. At 64 microM Cu2+, increases of necrosis up to 18% were seen whereas comparable levels of apoptotic and necrotic cells (<5%) were seen below this concentration. Chronic exposure over 8 weeks impaired colony-forming efficiency at concentrations of 16 microM Cu2+ and above. This study suggests that when liver cells sequester large amounts of copper, the toxic effects include delayed cell-cycle progression, a gradual loss of replicative capacity, and an increased incidence of cell death.

Background & Aims: The expression of matrix metalloproteinases 1, 2, 3, and 9 was examined in biopsy specimens removed from adult and pediatric patients with ulcerative colitis and Crohn's disease. The aim of this study was to determine if the expression of these enzymes was altered between areas of actively inflamed vs. noninvolved mucosa in the same patient and between patients with diseased bowel vs. a control group of patients., Methods: Proteolytic activity was quantified by zymography using image analysis. The identity of the matrix metalloproteinases was confirmed by using inhibitors, by comparison with purified standards, and by Western immunoblotting with specific antibodies., Results: In patients with ulcerative colitis (n = 21), a significant increase (P = 0.0051) in metalloproteinase activity was found in inflamed areas of mucosa compared with noninvolved regions. The levels of activity also were significantly greater (P < 0.001) in noninvolved areas of the bowel (n = 21) compared with levels in control patients (n = 9). In Crohn's disease (n = 8), differences between ulcerated and nonulcerated sites were not significantly different but levels of protease activity at both of these sites were significantly elevated compared with levels in control patients (P < 0.03). Of the proteases detected, matrix metalloproteinase 9 was the most abundantly expressed in the inflamed bowel; neutrophils were confirmed as the likely origin of this protease., Conclusions: The abundance and activation of matrix metalloproteinases significantly increases in ulcerative colitis and Crohn's mucosa. Inhibitors of these proteolytic enzymes may therefore be of therapeutic value in the treatment of inflammatory bowel disease.

A peptide specific antibody (AH1OW1) was raised against an epitope, AH10 (aa 449-463), of the alpha1(IV) chain adjacent to a cleavage site for matrix metalloproteinases (MMP)-2 and -9 within the triple helix of type IV collagen. The antibody only reacted with denatured and reduced preparations of type IV collagen, or with pepsin isolated type IV collagen digested with MMP-2 and MMP-9. The specificity of this antibody for the denatured triple helix was demonstrated by the lack of staining with pre-immune antibody and by pre-incubation of AH1OW1 antibody with excess AH10 peptide epitope. The AH1OWI antibody was used to detect whether proteolysis of type IV collagen occurs in ulcerative colitis, an inflammatory bowel condition often characterised by a large influx of granulocytes and macrophages and an associated tissue destruction. However, no evidence of in situ proteolysis of the basement membrane type IV collagen was observed. Only in the most actively inflamed mucosa was staining with AH1OW1 antibody observed in the mucosal connective tissue. Digestion of frozen sections of bowel with MMP-1, MMP-2, MMP-3 and MMP-9 did not result in the exposure of the AH10 epitope. These data demonstrate the stability of intact type IV collagen and indicate that susceptibility of alpha1(IV) chain to digestion with MMP-2 and MMP-9 may require other proteolytic/denaturing events in the molecule.

The mouse Cdx1 gene encodes a homeobox-containing transcription factor and is one of the few homeobox genes known to be expressed in endodermally derived tissues of the intestine in fetal and adult mice. A detailed and systematic study of the expression of the Cdx1 protein was carried out during embryonic intestinal development, postnatal cytodifferentiation and in the regenerating (after radiation-induced damage) intestine of the mouse. Using antibodies directed against Cdx1, we show for the first time that the Cdx1 protein is localised in the proliferating immature epithelium during intestinal development. It becomes restricted to the proliferative crypt compartment during postnatal differentiation, as well as in the adult intestine. The mesenchymal layer was completely negative both during embryonic development and in the postnatal intestine. The expression of the protein was first clearly detected throughout the simple columnar epithelium at day 15 of development. This expression progressively became restricted to the regions of epithelial proliferation in the crypts of the adult mouse by day 40 of post-natal development. There were occasional cells that were Cdx1 positive in the villi. During regeneration of the epithelium after radiation-induced damage, Cdx1 expression diminished during the initial phase of cellular regression. The expression was then very strong in the regenerating epithelial foci, but not in the quiescent sterilised crypts between day 4 and 6. The normal pattern was restored between day 6 and 7. The Paneth cells were negative. The physical segregation of Cdx1 with the proliferative compartment and the hierarchy of cell renewal in the intestinal epithelium is an important example of how regulatory genes function in the maintenance and in the dysfunction of renewing tissues.

Pneumonia caused by Streptococcus pneumoniae is characterized by neutrophil infiltration and variable epithelial injury. Neutrophil adhesion to alveolar epithelial pneumocytes (A549) was measured and demonstrated to be dose-dependent following preincubation of these (A549) pneumocytes with type 1 S. pneumoniae. Adhesion peaked at a bacteria-to-epithelial cell ratio of 5:1 after a 4-h incubation but was absent after 2 h and without FMLP. Filtered conditioned media (CM) from pneumococci cultured with (CM+) or without (CM-) epithelial cells were tested. CM+ induced significant adhesion in the absence of FMLP (P < .001); CM- had no effect. In the presence of FMLP, adhesion induced by both media was significantly greater than by FMLP alone (P < .001) and was significantly blocked (P < .01) by antibodies to CD11b and CD18. CM+ upregulated epithelial intercellular adhesion molecule 1 but CM- did not. These data provide new information concerning the interactions of S. pneumoniae, alveolar epithelial cells, and neutrophils.

Background and Aims: Conventional models of postnatal mucosal regeneration are cumbersome and limited: a novel model is described here. In addition, the influence of cell interactions on mucosal regeneration is examined within the model., Methods: Postnatal rat small intestinal mucosa was digested by enzymes to yield heterotypic cell aggregates (CA). CA colony forming ability, growth, and limited cytodifferentiation were assessed in vitro. CA were transplanted subcutaneously and retrieved for histological examination at staggered intervals to assess neomucosal morphogenesis and cytodifferentiation in vivo. Cell interactions in CA were disrupted by enzymes, thus producing cell suspensions (CS). Regeneration by CA and CS were compared., Results: CA produced proliferative colonies in vitro and showed a temporal sequence of neomucosal morphogenesis and differentiation in vivo. CA colonies were more numerous within 24 hours of primary culture and had greater cellularity by 96 hours than CS colonies. Alkaline phosphatase was expressed only by 258 of 696 CA colonies (37%). CA subcutaneous grafts (48 of 56 (87%)) regenerated small intestinal neomucosa while CS were unsuccessful., Conclusion: These methods provide a model of mucosal regeneration which includes constituent processes of colony formation, growth, neomucosal morphogenesis, and cytodifferentiation. Preservation of cell interactions within CA seems advantageous to regeneration within the model.

The 5T4 oncofoetal antigen is a 72 kDa glycoprotein defined by a monoclonal antibody raised against human placental trophoblast and is expressed in many different carcinomas but detected only at low levels in some normal epithelia. Immunohistochemical analysis of the patterns of expression in colorectal carcinomas has indicated a significant association between the presence of the antigen in tumour cells and metastatic spread. A cDNA encoding the 5T4 molecules has been isolated and the extracellular portion contains several leucine-rich repeats which have been implicated in cellular interactions. To study the cell biological role of 5T4 molecules, murine L cells (A9 derivative) were stably transfected with 5T4 cDNA under the control of the CMV immediate-early promoter. The 5T4-expressing cells exhibited a more spindle-shaped morphology compared to the vector alone transfected cells. Confocal immunofluorescence microscopy revealed a 'polkadot' pattern of 5T4 antigen expression, heterogeneous in intensity between cells, but distributed over the entire cell surface. Transmission and scanning electron microscopy showed that the 5T4 antigen is concentrated at microvillus projections of the plasma membrane both in the transfected A9 cells and in various carcinoma cell lines. Such projections express an array of surface molecules which function in cell adhesion and motility. This association of 5T4 antigen with microvillus projections was also observed in various carcinoma cell lines. 5T4 expression in A9 cells was also associated with an altered pattern of cell division, decreased cell-substratum adhesion and increased cellular motility. These results support the hypothesis that 5T4 molecules may have a direct function in trophoblast and tumour cell invasion processes.

Although the intestinal epithelium is one of the most rapidly renewing tissues, little is known about the major growth factors that control the rate of cell replacement and migration. Recently, a primary culture model has been described for the developing rat small intestinal epithelium, which permits epithelial growth while maintaining interactions with associated stromal cells, thereby possessing several contextual advantages over established cell lines (Evans et al., 1992). We have used this model to begin to determine the factors that may be involved in controlling intestinal epithelial cell proliferation. Under the conditions examined, no single growth factor promoted exclusive proliferation of epithelial cells; stromal cell proliferation was also apparent. The most potent stimulators of epithelial proliferation were insulin and insulin-like growth factor 1 (IGF-1). These factors also appeared to inhibit migration of the epithelial cells. 5-10 ng/ml EGF, 5-20 ng/ml TGF alpha, and 10-20 ng/ml PDGF also slightly increased epithelial cell numbers. Cell proliferation was inhibited by 0.1 ng/ml TGF beta-1. In Dulbecco's modified Eagle's medium (DMEM) containing 0.25 IU/ml insulin, glucose levels of 2-3 g/liter permitted epithelial growth with limited expansion of the stromal cell population. Higher levels of glucose further stimulated the nonepithelial cell types. Transferrin was also a potent stimulator of both cell types.

Basic fibroblast growth factor (bFGF) is found in the extracellular matrix and around the endothelial and epithelial cells of some human colon carcinomas. It is believed to play a role in angiogenesis, but in addition, recent data suggest that it can directly stimulate mitogenesis in some colon carcinoma cell lines. To clarify the role of bFGF in human colon carcinoma, we developed a model of Caco-2 which grew in serum-free conditions so that the effect of bFGF on multiplication, migration, and differentiation could be studied in defined conditions. Through morphological and biochemical studies in serum-free conditions, we demonstrated that this subline of Caco-2 differentiated spontaneously on reaching confluence. Using this model, we found that bFGF did not affect differentiation but that multiplication and migration were increased. The implication of these findings is that bFGF, released from the extracellular matrix by invading cells or produced by neovascular endothelial cells, can increase the mitogenic rate and migratory potential of colon carcinoma cells. In addition, the dual role of bFGF in stimulating colon carcinoma cells directly and promoting angiogenesis suggests that anti-bFGF strategies could form the basis of a novel approach to the treatment of colon carcinoma.

A novel method to study the generation of rat small intestinal mucosa, by transplantation of disaggregated postnatal rat small intestinal epithelium is described. Cellular aggregates, comprised of epithelium with attached proliferative cells and closely associated stromal tissue, were isolated from postnatal rat small intestine by enzymatic digestion, then grafted immediately to the subcutaneous plane of adult recipients. On graft retrieval after 14 days, 39% of cellular transplants to nude mice, and 84% of cellular transplants to inbred rats had developed into small intestine-like structures. These structures were comprised of a circumferential layer of epithelium surrounding a central mucin filled lumen. This neomucosal layer exhibited well formed crypts and villi, and contained all epithelial stem cell lineages i.e. absorptive enterocytes, goblet cells, Paneth's cells and entero-endocrine cells. Proliferative activity within this neomucosa was confined to crypt regions as in normal postnatal small intestine. Developmental maturation within the regenerated neomucosa was demonstrated by organotypic morphogenesis, i.e. formation of mature crypts and villi, and progressive cytodifferentiation with increased numbers of goblet cells, entero-endocrine cells and Paneth's cells. Altered patterns of brush border enzyme expression further confirmed a temporal progression of development within neomucosal enterocytes. It is concluded that after "extensive" mucosal disaggregation, postnatal small intestinal epithelial progenitor cells retain the capacity for organotypic regeneration of neomucosa when transplanted to ectopic sites in adult recipients. These small aggregates of epithelium and stroma are capable of generating the topographical signals necessary for the three dimensional regeneration of this tissue. Furthermore, the multipotent generative potential of the stem cells within these cellular aggregates is maintained with production of all progeny.

Heparin is a sulphated glycosaminoglycan derived from mast cells and has a number of functions including the inhibition of proliferation in several cell types and interactions with a range of heparin-binding growth factors. We report that heparin is a trophic factor in primary cultures of rat small intestinal epithelium. Heparin elicits a dose-dependent increase in epithelial proliferation and inhibits the growth of associated mesenchyme. The trophic effect of this molecule is not reproduced by other glycosaminoglycans including heparan sulphate but is dependent upon extensive molecular sulphation. Highly sulphated polysaccharides that are structurally unrelated to heparin (e.g. dextran sulphate and pentosan polysulphate) also stimulate epithelial proliferation in primary cultures. Heparin may act by the potentiation of mesenchyme-derived heparin-binding growth factors and these data suggest an in vivo role for mast cell-derived heparin in mucosal wound regeneration.