Translational research into the effects of cigarette smoke on inflammatory mediators and epithelial TRPV1 in Crohn's disease.

Crohn's disease is a pathological condition of the gastro-intestinal tract, causing severe transmural inflammation in the ileum and/or colon. Cigarette smoking is one of the best known environmental risk factors for the development of Crohn's disease. Nevertheless, very little is known about the effect of prolonged cigarette smoke exposure on inflammatory modulators in the gut. We examined the effect of cigarette smoke on cytokine profiles in the healthy and inflamed gut of human subjects and in the trinitrobenzene sulphonic acid mouse model, which mimics distal Crohn-like colitis. In addition, the effect of cigarette smoke on epithelial expression of transient receptor potential channels and their concurrent increase with cigarette smoke-augmented cytokine production was investigated. Active smoking was associated with increased IL-8 transcription in ileum of controls (p < 0,001; n = 18-20/group). In the ileum, TRPV1 mRNA levels were decreased in never smoking Crohn's disease patients compared to healthy subjects (p <0,001; n = 20/group). In the colon, TRPV1 mRNA levels were decreased (p = 0,046) in smoking healthy controls (n = 20/group). Likewise, healthy mice chronically exposed to cigarette smoke (n = 10/group) showed elevated ileal Cxcl2 (p = 0,0075) and colonic Kc mRNA levels (p = 0,0186), whereas TRPV1 mRNA and protein levels were elevated in the ileum (p = 0,0315). Although cigarette smoke exposure prior to trinitrobenzene sulphonic acid administration did not alter disease activity, increased pro-inflammatory cytokine production was observed in the distal colon (Kc: p = 0,0273; Cxcl2: p = 0,104; Il1-ß: p = 0,0796), in parallel with the increase of Trpv1 mRNA (p < 0,001). We infer that CS affects pro-inflammatory cytokine expression in healthy and inflamed gut, and that the simultaneous modulation of TRPV1 may point to a potential involvement of TRPV1 in cigarette smoke-induced production of inflammatory mediators.

Transient Receptor Potential Channels in Intestinal Inflammation: What Is the Impact of Cigarette Smoking?

Inflammatory bowel disease (IBD) is characterized by severe gastrointestinal inflammation and results from a complex interplay between genetic and environmental factors. IBD includes two prominent subtypes: Crohn's disease (CD) and ulcerative colitis (UC). One of the main risk factors for the development of CD is cigarette smoking, while UC is rather a disease of ex-smokers. To date, many of the mechanisms underlying the immune imbalance in IBD and the involvement of cigarette smoke (CS) are incompletely understood. Transient receptor potential (TRP) proteins are non-selective cation channels that, upon activation, lead to plasma membrane depolarization and, in general, to Ca2+ influx. TRP channels of the ankyrin and vanilloid family, expressed by sensory neurons in the central and enteric nervous systems, have been extensively studied in the context of intestinal inflammation. Moreover, recent advances made on the role of non-neuronal expressed TRP channels shed light on the involvement of epithelial cells in inflammatory processes. This review focuses on how CS may impact TRP channel function in intestinal inflammation. Firstly, we discuss the current knowledge on neuronal TRP channels, known to be linked to IBD, in health, immune homeostasis and intestinal inflammation. Subsequently, we address how TRP channels are activated by CS and its components in other organ systems and also hypothesize on the potential implications for CS-mediated TRP channel activation in gut inflammation.

Straightforward and sensitive RT-qPCR based gene expression analysis of FFPE samples.

Fragmented RNA from formalin-fixed paraffin-embedded (FFPE) tissue is a known obstacle to gene expression analysis. In this study, the impact of RNA integrity, gene-specific reverse transcription and targeted cDNA preamplification was quantified in terms of reverse transcription polymerase chain reaction (RT-qPCR) sensitivity by measuring 48 protein coding genes on eight duplicate cultured cancer cell pellet FFPE samples and twenty cancer tissue FFPE samples. More intact RNA modestly increased gene detection sensitivity by 1.6 fold (earlier detection by 0.7 PCR cycles, 95% CI = 0.593-0.850). Application of gene-specific priming instead of whole transcriptome priming during reverse transcription further improved RT-qPCR sensitivity by a considerable 4.0 fold increase (earlier detection by 2.0 PCR cycles, 95% CI = 1.73-2.32). Targeted cDNA preamplification resulted in the strongest increase of RT-qPCR sensitivity and enabled earlier detection by an average of 172.4 fold (7.43 PCR cycles, 95% CI = 6.83-7.05). We conclude that gene-specific reverse transcription and targeted cDNA preamplification are adequate methods for accurate and sensitive RT-qPCR based gene expression analysis of FFPE material. The presented methods do not involve expensive or complex procedures and can be easily implemented in any routine RT-qPCR practice.

Chronic cigarette smoke exposure induces microbial and inflammatory shifts and mucin changes in the murine gut.

Inflammatory bowel diseases (IBD) are complex multifactorial diseases characterized by an inappropriate host response to an altered commensal microbiome and dysfunctional mucus barrier. Cigarette smoking is the best known environmental risk factor in IBD. Here, we studied the influence of chronic smoke exposure on the gut microbiome, mucus layer composition and immune factors in conventional mice. We compared smoke-exposed with air-exposed mice (n = 12) after a smoke exposure of 24 weeks. Both Illumina sequencing (n = 6) and denaturing gradient gel electrophoresis (n = 12) showed that bacterial activity and community structure were significantly altered in the colon due to smoke exposure. Interestingly, an increase of Lachnospiraceae sp. activity in the colon was observed. Also, the mRNA expression of Muc2 and Muc3 increased in the ileum, whereas Muc4 increased in the distal colon of smoke-exposed mice (n = 6). Furthermore, we observed increased Cxcl2 and decreased Ifn-γ in the ileum, and increased Il-6 and decreased Tgf-ß in the proximal colon. Tight junction gene expression remained unchanged. We infer that the modulating role of chronic smoke exposure as a latently present risk factor in the gut may be driven by the altered epithelial mucus profiles and changes in microbiome composition and immune factors.

The Effect of Cigarette Smoke Exposure on the Development of Inflammation in Lungs, Gut and Joints of TNFΔARE Mice.

The inflammatory cytokine TNF-α is a central mediator in many immune-mediated diseases, such as Crohn's disease (CD), spondyloarthritis (SpA) and chronic obstructive pulmonary disease (COPD). Epidemiologic studies have shown that cigarette smoking (CS) is a prominent common risk factor in these TNF-dependent diseases. We exposed TNFΔARE mice; in which a systemic TNF-α overexpression leads to the development of inflammation; to 2 or 4 weeks of air or CS. We investigated the effect of deregulated TNF expression on CS-induced pulmonary inflammation and the effect of CS exposure on the initiation and progression of gut and joint inflammation. Upon 2 weeks of CS exposure, inflammation in lungs of TNFΔARE mice was significantly aggravated. However, upon 4 weeks of CS-exposure, this aggravation was no longer observed. TNFΔARE mice have no increases in CD4+ and CD8+ T cells and a diminished neutrophil response in the lungs after 4 weeks of CS exposure. In the gut and joints of TNFΔARE mice, 2 or 4 weeks of CS exposure did not modulate the development of inflammation. In conclusion, CS exposure does not modulate gut and joint inflammation in TNFΔARE mice. The lung responses towards CS in TNFΔARE mice however depend on the duration of CS exposure.

Pathologic Assessment of Rectal Carcinoma after Neoadjuvant Radio(chemo)therapy: Prognostic Implications.

Neoadjuvant radio(chemo)therapy is increasingly used in rectal cancer and induces a number of morphologic changes that affect prognostication after curative surgery, thereby creating new challenges for surgical pathologists, particularly in evaluating morphologic changes and tumour response to preoperative treatment. Surgical pathologists play an important role in determining the many facets of rectal carcinoma patient care after neoadjuvant treatment. These range from proper handling of macroscopic specimens to accurate microscopic evaluation of pathological features associated with patients' prognosis. This review presents the well-established pathological prognostic indicators and discusses challenging features in order to provide both surgical pathologists and treating physicians with a checklist that is useful in a neoadjuvant setting.

Determinants of testosterone levels in human male obesity.

Testosterone (T) levels are decreased in obese men, but the underlying causes are incompletely understood. Our objective was to explore the relation between low (free) T levels and male obesity, by evaluating metabolic parameters, subcutaneous adipose tissue (SAT) aromatase expression, and parameters of the hypothalamic-pituitary-gonadal axis. We recruited 57 morbidly obese men [33 had type 2 diabetes (DM2)] and 25 normal-weight men undergoing abdominal surgery. Fourteen obese men also attended a follow-up, 2 years after gastric bypass surgery (GBS). Circulating T levels were quantified by LC-MS/MS, whereas free T levels were measured using serum equilibrium dialysis and sex hormone-binding globulin, luteinizing hormone, and follicle-stimulating hormone by immunoassay. SAT biopsies were used to determine adipocyte cell size and aromatase expression by real-time PCR. Total and free T levels were decreased in obese males versus controls, with a further decrease in obese men with DM2 versus obese men without DM2. There were no differences in aromatase expression among the study groups, and sex steroids did not correlate with aromatase expression. Pearson analysis revealed an inverse association between (free) T and SAT cell size, triglycerides, and HOMA-IR. Multivariate analysis confirmed the inverse association between (free) T and SAT cell size (ß = -0.321, P = 0.037 and ß = -0.441, P = 0.011, respectively), independent of age, triglycerides, HOMA-IR, obesity, or diabetes. T levels were normalized 2 years after GBS. These data suggest that SAT cell size rather than SAT aromatase expression or parameters of the hypothalamic-pituitary-gonadal axis is related to low T in male obesity, which points to adipose cell size-related metabolic changes as a major trigger in decreased T levels.

Commensal microbiota influence systemic autoimmune responses.

Antinuclear antibodies are a hallmark feature of generalized autoimmune diseases, including systemic lupus erythematosus and systemic sclerosis. However, the processes underlying the loss of tolerance against nuclear self-constituents remain largely unresolved. Using mice deficient in lymphotoxin and Hox11, we report that approximately 25% of mice lacking secondary lymphoid organs spontaneously develop specific antinuclear antibodies. Interestingly, we find this phenotype is not caused by a defect in central tolerance. Rather, cell-specific deletion and in vivo lymphotoxin blockade link these systemic autoimmune responses to the formation of gut-associated lymphoid tissue in the neonatal period of life. We further demonstrate antinuclear antibody production is influenced by the presence of commensal gut flora, in particular increased colonization with segmented filamentous bacteria, and IL-17 receptor signaling. Together, these data indicate that neonatal colonization of gut microbiota influences generalized autoimmunity in adult life.

Follicles of various maturation stages react differently to enzymatic isolation: a comparison of different isolation protocols.

Isolation of human follicles is based on digestion of the tissue by combinations of enzymes. Follicle vitality and morphology are often based on the analysis of pooled follicles of different maturation stages. Information is therefore lacking on the effect of the isolation protocol to individual follicles of different maturation stages. A study was conducted using five protocols combining different enzymes and varying concentrations. Isolated follicles were classified according to their maturation stages, counted and characterized for vitality, morphology, early apoptosis and organization of transzonal projections. No statistical differences were found between the protocols when outcome parameters were analysed on a pool of follicles regardless of their maturation status. Differences were observed in quality when the follicles were analysed separately according to their maturation status. Combining morphologic characteristics and vitality, both Liberase DH and Liberase TM combined with collagenase IV were better at isolating high-quality primordial follicles, compared with collagenase IV. No statistical difference between the isolation protocols was found for primary follicles. If only high-quality isolated secondary follicles are needed, collagenase IV is found to be most advantageous. Follicles of different maturation stages react differently when enzymatic isolation protocols are compared.

Spray-dried polyelectrolyte microparticles in oral antigen delivery: stability, biocompatibility, and cellular uptake.

During the past decade, extensive research has undeniably improved the formulation and delivery of oral vaccines. Nevertheless, several factors, such as the harsh gastrointestinal environment together with tolerance induction to exogenous antigens, have thus far impeded the optimal effectiveness and clinical application of oral delivery systems. The current study encompasses an initial evaluation of the stability, biocompatibility, and cellular uptake of two promising candidate systems for oral antigen delivery, that is, calcium carbonate- (CP) and mannitol-templated (MP) porous microspheres. Both spray-dried formulations were efficiently internalized by human intestinal epithelial cells (Caco-2 and HT-29) and degraded into phagolysosomal intracellular compartments. In addition, cellular particle uptake and processing significantly up-regulated the expression of (HLA) class-II and costimulatory molecules on intestinal epithelial cells. Even though the high surface-area-to-volume ratio of the microspheres was expected to favor protease access, antigen release was remarkably limited in simulated intestinal fluid and was even absent under gastric conditions. Finally, neither CP nor MP exerted cytotoxicity upon prolonged in vitro incubation with high antigen concentration. Altogether, these data support the potential of CP and MP for oral antigen delivery and motivate the further development of these promising carrier systems in in vivo studies.

Recent advances in oral vaccine development: yeast-derived ß-glucan particles.

Oral vaccination is the most challenging vaccination method due to the administration route. However, oral vaccination has socio-economic benefits and provides the possibility of stimulating both humoral and cellular immune responses at systemic and mucosal sites. Despite the advantages of oral vaccination, only a limited number of oral vaccines are currently approved for human use. During the last decade, extensive research regarding antigen-based oral vaccination methods have improved immunogenicity and induced desired immunological outcomes. Nevertheless, several factors such as the harsh gastro-intestinal environment and oral tolerance impede the clinical application of oral delivery systems. To date, human clinical trials investigating the efficacy of these systems are still lacking. This review addresses the rationale and key biological and physicochemical aspects of oral vaccine design and highlights the use of yeast-derived ß-glucan microparticles as an oral vaccine delivery platform.

ß-Glucan microparticles are good candidates for mucosal antigen delivery in oral vaccination.

Continuously improving the developmental process and the efficacy of oral vaccines is essential in the fight against intestinal pathogens. A promising strategy for vaccination applying safe, biodegradable and non-replicating antigen delivery systems has gained increased interest for eliciting cellular and humoral immune responses. The current study evaluates the potential of ß-glucan particles (GP) as an oral antigen delivery system and their adjuvant characteristics. GP are efficiently internalized by human intestinal epithelial cell lines (Caco-2 and HT-29 cells), without exerting negative effects on cell viability. GP triggered the expression of pro-inflammatory cytokines IL-23p19, IL-8 and the ß-glucan receptors dectin-1 and TLR2 by activated Caco-2 cells, and CCL20 in HT-29 cells. In contrast, the expression level of TGF-ß, an important mediator of oral tolerance, was significantly downregulated in HT-29 cells. Additionally, adoptive transfer experiments showed proliferating ovalbumin (OVA)-specific CD4(+) T cells mainly in the spleens of GP-OVA-fed mice. Furthermore, we detected a significantly increased IL-17 and a trend towards increased IFN-γ production in the spleen of GP-OVA-fed mice upon antigen restimulation. Oral administration of GP-OVA induced increased OVA-specific IgA, secretory-IgA (S-IgA) and secretory component (SC) production in intestinal fluids. Our data show that GP vehicles are able to deliver OVA via an oral route allowing efficient antigen presentation alongside adaptive immune activation, resulting in a Th17-biased response and the production of OVA-specific IgA, secretory-IgA and secretory component antibodies.

Inflammatory patterns in upper airway disease in the same geographical area may change over time.

BACKGROUND: Inflammatory patterns of nasal polyps (NPs) may vary. Changes over time have not been investigated so far. This study was designed to evaluate the inflammatory patterns of NPs in Thailand at two time points 12 years apart, explore differences in Staphylococcus aureus (SA) mucosal carriage rates over time, and the latter's relationship with the inflammatory patterns. METHODS: Formalin-fixed nasal tissue was obtained from 89 (47 in 1999 and 42 in 2011) patients suffering from chronic rhinosinusitis with NPs (CRSwNPs). Tissues were evaluated for eosinophils, neutrophils, IgE(+) cells, IgE and macrophage mannose receptors, interleukin (IL)-5 and IL-17 cytokine profile, and the presence of SA, using automated immunohistochemistry and peptide nucleic acid-fluorescence in situ hybridization. RESULTS: We found a significant increase in the absolute values of eosinophils and IgE(+) cells in the 2011 CRSwNP tissue series compared with 1999 and a significant but smaller increase in neutrophils. Semiquantitative evaluation revealed significantly higher mean values of positive cells for all studied inflammatory markers in the 2011 group of patients, except for the high-affinity IgE receptor. This "eosinophilic shift" of inflammation was accompanied by higher SA carriage, as well as higher frequencies of SA invasion (54.8% versus 10.6%; p < 0.001) in the 2011 compared with 1999 subjects. Patients with asthma were more likely to have higher SA carriage rates compared with nonasthmatic patients. CONCLUSION: There was a shift from predominantly neutrophilic to eosinophilic CRSwNPs in Thai patients within 12 years, with an increase in various inflammatory markers including IgE, which is associated with an increase in intramucosal presence of SA.

The critical apical diameter to obtain regeneration of the pulp tissue after tooth transplantation, replantation, or regenerative endodontic treatment.

INTRODUCTION: Regeneration of pulp-like tissue in the pulp chamber after tooth transplantation, replantation, or in regenerative endodontic treatment is only possible if the apical foramen is open. According to the literature, the success of regeneration decreases considerably if the foramen is smaller than 1 mm when measured on radiographs. The aim of this study was to study histologically the relation between the width of the apical foramen and regeneration of tissue in the pulp chamber after autotransplantation. METHODS: Fifteen single-rooted mature teeth of 3 adult beagle dogs were used. All experimental teeth were extracted and underwent apicoectomy. The teeth were photographed from the apical side, and the width of the foramen was calculated. The foramen width ranged from 0.24-1.09 mm. All teeth were replanted in infraocclusion. The observation period was 90 days after transplantation. RESULTS: The 10 teeth with the smallest apical diameter, ranging between 0.24 and 0.53 mm, showed vital tissue in at least one third of the pulp chamber. The 6 most successful teeth showing vital tissue in the entire pulp chamber had an apical diameter between 0.32 and 0.65 mm, and 80% of the experimental teeth with a diameter varying between 1.09 and 0.31 mm showed vital tissue in at least one third of the pulp chamber 90 days after transplantation. CONCLUSIONS: The size of the apical foramen seems not to be the all decisive factor for successful revascularization and ingrowth of new tissue after transplantation. The minimum width of the apical foramen has not been determined, but a size smaller than 1 mm does not prevent revascularization and ingrowth of vital tissue. In this animal study an apical foramen of 0.32 mm did not prevent ingrowth of new tissue in two-thirds of the pulp chamber 90 days after transplantation.

Different types of tissue composition in inflammatory or reparative upper airway disorders.

BACKGROUND: Composition changes of extracellular matrix (ECM) can lead to functional disorders of the upper airways (UA). The aim of this study was to systematically measure both the association patterns and the correlation degree between tissue composition parameters in UA inflammatory diseases. METHODOLOGY: Nasal samples were obtained from patients with chronic rhinosinusitis with (CRS+NP), without nasal polyps (CRS), with post-operative adhesions (S) and normal nasal mucosa (NM). A reproducible semi-quantitative method, which takes epithelial and lamina propria damages into account was applied for haematoxylin and eosin, alpha-smooth muscle actin, reticulin, elastin, laminin and collagen type IV stainings. RESULTS: The most severe cases of epithelial shedding have been found in a significant higher amount in CRS+NP when compared with NM. The most severe cases of inflammatory reaction were mainly found in CRS+NP. CRS+NP had significantly more severe cases of oedema than NM. Excluding elastin, networks in other ECM proteins were found modified in fibrotic fields but to a lesser extend in oedematous regions in all conditions. CONCLUSION: Although non specific, oedema in the lamina propria is a key-feature of CRS+NP, while fibrosis, massively present in CRS and S, affects profoundly the distribution of ECM proteins in these areas.

The effect of smoking on intestinal inflammation: what can be learned from animal models?

Epidemiological evidence demonstrates that smoking is the most important environmental risk factor in Crohn's disease while it positively interferes with the disease course of ulcerative colitis. However, the underlying mechanisms through which smoking exerts this divergent effect and affects pathogenesis of inflammatory bowel disease are largely unknown. Animal smoke models are good models to investigate the impact of cigarette smoke on intestinal physiology and inflammation. They enable one to explore the interaction of smoke components and the gut on cellular and molecular level, clarifying how smoking interferes with normal gut function and with disease course in inflammatory conditions. This review describes the currently used animal models for studying the impact of cigarette smoke on the intestinal tract. We first discuss the different methods for simulation of smoking. Furthermore, we focus on the effect of smoke exposure on normal gut physiology and immunology, on experimental (entero)colitis, and on inflammation-induced neoplasia. Based on this current knowledge, a hypothesis is formulated about the mechanisms through which cigarette smoke interferes with the gut in normal and pathological conditions.

Cigarette smoke and the terminal ileum: increased autophagy in murine follicle-associated epithelium and Peyer's patches.

Cigarette smoke (CS) exposure is associated with increased autophagy in several cell types, such as bronchial epithelial cells. Smoking is also an environmental risk factor in Crohn's disease, in which impairment of the autophagy-mediated anti-bacterial pathway has been implicated. So far, it is unknown whether CS induces autophagy in the gut. Here, we examined the effect of chronic CS exposure on autophagy in the follicle-associated epithelium (FAE) of murine Peyer's patches. Transmission electron microscopy revealed that the proportion of cell area occupied by autophagic vesicles significantly increased in the FAE after CS exposure. An increased number of autophagic vesicles was observed in the FAE, whereas the vesicle size remained unaltered. Besides enterocytes, also M-cells contain more autophagic vesicles upon CS exposure. In addition, the mRNA level of the autophagy-related protein Atg7 in the underlying Peyer's patches is increased after CS exposure, which indicates that the autophagy-inducing effect of CS is not limited to the FAE. In conclusion, our results demonstrate that CS exposure induces autophagy in murine FAE and in the underlying immune cells of Peyer's patches, suggesting that CS exposure increases the risk for Crohn's disease by causing epithelial oxidative damage, which needs to be repaired by autophagy.

Cigarette smoking alters epithelial apoptosis and immune composition in murine GALT.

Smokers have a twofold increased risk to develop Crohn's disease (CD). However, little is known about the mechanisms through which smoking affects CD pathogenesis. Especially Crohn's ileitis is negatively influenced by smoking. Interestingly, the ileum and, more in particular, the Peyer's patches in the terminal ileum are also the sites where the first CD lesions are found. Several chemokines are implicated in the pathogenesis, among which is the CCL20-CCR6 pathway. Here, we studied the gut-associated lymphoid tissue in C57BL/6 wild-type mice and in CCR6-deficient mice after exposure to air or cigarette smoke for 24 weeks. Apoptotic index of the follicle-associated epithelium overlying the Peyer's patches was evaluated. We found that chronic smoke exposure induced apoptosis in the follicle-associated epithelium. Furthermore, immune cell numbers and differentiation along with chemokine expression were determined in Peyer's patches. Important changes in immune cell composition were observed: total dendritic cells, CD4+ T cells (including regulatory T cells) and CD8+ T cells increased significantly after smoke exposure. The CD11b+ dendritic cell subset almost doubled. Interestingly, these changes were accompanied by an upregulated mRNA expression of the chemokines CCL9 and CCL20. However, no differences in the increase of dendritic cells were observed between wild-type and CCR6-deficient mice. Our results show that cigarette smoke exposure increases apoptosis in the follicle-associated epithelium and is associated with immune cell accumulation in Peyer's patches.

Does removal of the original pulp tissue before autotransplantation influence ingrowth of new tissue in the pulp chamber?

In an attempt to extend the indication area for autotransplantation of vital teeth, two possibilities can be proposed: (i) The enlargement of the apical foramen, with the aim to facilitate revascularization and ingrowth of new tissue. The ingrowth of tissue will eliminate the need for endodontic treatment when mature teeth are transplanted and (ii) the cryopreservation of teeth in case they cannot be transplanted immediately to the receptor site. Teeth with an ideal stage of root formation can be cryopreserved to perform transplantation later. Although pulpcell cultures survive crypreservation in vitro, the pulp tissue cannot survive the cryopreservation procedures when it is kept inside the pulpchamber. Therefore, the pulp tissue has to be removed before cryopreservation. It has been demonstrated that revascularization and ingrowth of new tissue can occur in an empty pulp chamber (1). The aim of this study was to find out if revascularization and ingrowth of new pulp tissue is influenced by removal of the original pulp tissue before autotransplantation. Twenty nine single-rooted teeth from three adult beagle dogs were transplanted after resection of the root tip. One group of teeth (n = 14) had the pulp tissue removed before transplantation. The other group (n = 15) had the original pulp left in situ. The transplanted teeth were histologically analysed 90 days post-transplantation. In the group with the tissue left in situ, 12 teeth (80%) showed a pulp chamber totally filled or at least 1/3 to 2/3 filled with viable tissue. In the group with the pulp tissue removed, 11 teeth (79%) had no or little vital tissue in the pulp chamber. The necrotic masses that develop in the original pulp tissue immediately after transplantation are a possible stimulating factor in the repair process of the pulp. As a conclusion, it can be stated that in case of autotransplantation of teeth, it is advisable to leave the pulp tissue in situ to stimulate the revascularization and ingrowth of new tissue after transplantation.

Identification of the nasal mucosa as a new target for leptin action.

UNLABELLED: The aim of this study was to examine systemic and local nasal leptin and leptin receptor expression in patients with nasal polyposis and healthy controls. METHODS AND RESULTS: Serum leptin and soluble leptin receptor levels were examined by enzyme-linked immunosorbent assay (ELISA). The presence of leptin and leptin receptor mRNA was investigated using reverse transcriptase-polymerase chain reaction (RT-PCR), and tissue leptin and leptin receptor protein expression was analysed by immunohistochemistry and ELISA. Serum levels of biologically active leptin were significantly elevated in patients with nasal polyps compared with control subjects. These serum leptin levels were strongly correlated with the levels found in tissue in both study groups, although leptin was not significantly elevated in nasal polyp tissue. Using RT-PCR, we showed that both leptin and its receptors were produced in nasal mucosa. Finally, immunohistochemistry showed that leptin and leptin receptor protein were expressed in several cells of the normal and inflamed nasal mucosa. CONCLUSIONS: Leptin receptors and their biological ligand leptin are expressed in the nasal mucosa, suggesting a possible role in upper airway immunology.