Biomedical subjects
Alessio Fasano
Publications and source records attributed to Alessio Fasano.
Permeability, zonulin production, and enteropathy in dermatitis herpetiformis.
BACKGROUND & AIMS: Dermatitis herpetiformis (DH) is characterized by variable degrees of enteropathy and increased intestinal permeability. Zonulin, a regulator of tight junctions, seems to play a key role in the altered intestinal permeability that characterizes the early phase of celiac disease. Our aim was to assess both intestinal permeability and serum zonulin levels in a group of patients with DH having variable grades of enteropathy. METHODS: We studied 18 DH patients diagnosed on the basis of characteristic immunoglobulin (Ig)A granular deposits in the dermal papillae of noninvolved skin. Results were compared with those of classic celiac patients, patients with linear IgA dermatosis, and healthy controls. RESULTS: According to Marsh's classification, 5 patients had no evidence of enteropathy (type 0), 4 patients had type II, 2 patients had type IIIb damage, and 7 patients had a more severe lesion (type IIIc). Intestinal permeability (lactulose/mannitol ratio [lac/man]) was abnormal in all patients with DH. Patients with more severe enteropathy had significantly greater permeability ( P < .05). The serum zonulin concentration (enzyme-linked immunosorbent assay) for patients with DH was 2.1 +/- .3 ng/mg with 14 of 16 (87.5%) patients having abnormally increased values. In contrast, patients with linear IgA dermatosis had normal histology, normal intestinal permeability, and negative celiac serology. CONCLUSIONS: Increased intestinal permeability and zonulin up-regulation are common and concomitant findings among patients with DH, likely involved in pathogenesis. Increased permeability can be observed even in patients with no evidence of histologic damage in biopsy specimens. Patients with linear IgA dermatosis appear to be a distinct population with no evidence of gluten sensitivity.
Mechanisms of disease: the role of intestinal barrier function in the pathogenesis of gastrointestinal autoimmune diseases.
The primary functions of the gastrointestinal tract have traditionally been perceived to be limited to the digestion and absorption of nutrients and electrolytes, and to water homeostasis. A more attentive analysis of the anatomic and functional arrangement of the gastrointestinal tract, however, suggests that another extremely important function of this organ is its ability to regulate the trafficking of macromolecules between the environment and the host through a barrier mechanism. Together with the gut-associated lymphoid tissue and the neuroendocrine network, the intestinal epithelial barrier, with its intercellular tight junctions, controls the equilibrium between tolerance and immunity to nonself-antigens. When the finely tuned trafficking of macromolecules is dysregulated in genetically susceptible individuals, both intestinal and extraintestinal autoimmune disorders can occur. This new paradigm subverts traditional theories underlying the development of autoimmunity, which are based on molecular mimicry and/or the bystander effect, and suggests that the autoimmune process can be arrested if the interplay between genes and environmental triggers is prevented by re-establishing intestinal barrier function. Understanding the role of the intestinal barrier in the pathogenesis of gastrointestinal disease is an area of translational research that encompasses many fields and is currently receiving a great deal of attention. This review is timely given the increased interest in the role of a 'leaky gut' in the pathogenesis of gastrointestinal diseases and the advent of novel treatment strategies, such as the use of probiotics.
Clinical presentation of celiac disease in the pediatric population.
Celiac disease (CD) is an immune-mediated enteropathy triggered by the ingestion of gluten-containing grains (including wheat, rye, and barley) in genetically susceptible individuals. CD is associated with HLA molecules DQ2 (90%-95%) and DQ8 (5%-10%), and in the continued presence of gluten the disease is self-perpetuating. CD is one of the most common lifelong disorders worldwide and is characterized by a variety of clinical presentations. These include the typical malabsorption syndrome (classic symptoms) and a spectrum of symptoms potentially affecting any organ or body system (nonclassic symptoms). Because CD often is atypical or even clinically silent, many cases go undiagnosed and are exposed to the risk of long-term complications. There is growing interest in the social aspects of CD because the burden of illness related to this condition is doubtless higher than previously thought.
Guideline for the diagnosis and treatment of celiac disease in children: recommendations of the North American Society for Pediatric Gastroenterology, Hepatology and Nutrition.
Celiac disease is an immune-mediated enteropathy caused by a permanent sensitivity to gluten in genetically susceptible individuals. It occurs in children and adolescents with gastrointestinal symptoms, dermatitis herpetiformis, dental enamel defects, osteoporosis, short stature, delayed puberty and persistent iron deficiency anemia and in asymptomatic individuals with type 1 diabetes, Down syndrome, Turner syndrome, Williams syndrome, selective immunoglobulin (Ig)A deficiency and first degree relatives of individuals with celiac disease. The Celiac Disease Guideline Committee of the North American Society for Pediatric Gastroenterology, Hepatology and Nutrition has formulated a clinical practice guideline for the diagnosis and treatment of pediatric celiac disease based on an integration of a systematic review of the medical literature combined with expert opinion. The Committee examined the indications for testing, the value of serological tests, human leukocyte antigen (HLA) typing and histopathology and the treatment and monitoring of children with celiac disease. It is recommended that children and adolescents with symptoms of celiac disease or an increased risk for celiac disease have a blood test for antibody to tissue transglutaminase (TTG), that those with an elevated TTG be referred to a pediatric gastroenterologist for an intestinal biopsy and that those with the characteristics of celiac disease on intestinal histopathology be treated with a strict gluten-free diet. This document represents the official recommendations of the North American Society for Pediatric Gastroenterology, Hepatology and Nutrition on the diagnosis and treatment of celiac disease in children and adolescents.
The impact of DeltaG on the oral bioavailability of low bioavailable therapeutic agents.
Low oral bioavailability continues to drive research toward identifying novel approaches to enhance drug delivery. Over the past few years, emphasis on the use of absorption enhancers has been overwhelming despite their major adverse effects. Zonula occludens toxin (Zot) was recently established as a safe and effective absorption enhancer, reversibly opening the tight junctions for hydrophilic markers and hydrophobic drugs across the small intestine and the blood brain barrier. DeltaG, the biologically active fragment of Zot, was isolated and shown to increase the in vitro transport and in vivo absorption of paracellular markers. The objective of this study was to examine the effect of DeltaG on the oral bioavailability of low bioavailable therapeutic agents. Jugular vein cannulated Sprague-Dawley rats were randomly assigned to receive the following treatments intraduodenally (ID): [(3)H]cyclosporin A, [(3)H]ritonavir, [(3)H]saquinavir, or [(3)H]acyclovir at (120 microCi/kg) alone, with protease inhibitors (PIs), or with DeltaG (720 microg/kg)/PI. Serial blood samples were collected, and plasma was analyzed for radioactivity. After ID administration with DeltaG/PI, C(max) significantly (p < 0.05) increased over a range of 197 to 5700%, whereas area under the plasma concentration time curve displayed significant increases extending over a range of 123.8 to 4990.3% for the investigated drugs. DeltaG significantly increased the in vivo oral absorption of some low bioavailable drugs in the presence of PI. This study suggests that DeltaG-mediated tight junction modulation, combined with metabolic protection, may be used to enhance the low oral bioavailability of certain drugs when administered concurrently.
Intestinal epithelial tight junctions as targets for enteric bacteria-derived toxins.
The application of a multidisciplinary approach to study bacterial pathogenesis, along with the recent sequencing of entire microbial genomes have made possible discoveries that are changing the way scientists view the bacterium-host interaction. Today, research on the molecular basis of the pathogenesis of infectious diarrheal diseases of necessity transcends established boundaries between microbiology, cell biology, intestinal pathophysiology, and immunology. Novel multidisciplinary approaches led to the discovery of new bacteria-host cell interactions involving signals regulating intestinal permeability through the modulation of cell cytoskeleton and intercellular tight junctions (TJ). A century ago, TJ were conceptualized as a secreted extracellular cement forming an absolute and unregulated barrier within the paracellular space. Biological studies of the past several decades have shown that TJ are dynamic structures subjected to structural changes that dictate their functional status under a variety of developmental, physiological, and pathological circumstances. To meet the many diverse physiological challenges to which the intestinal epithelial barrier is subjected, TJ must be capable of rapid and coordinated responses. This requires the presence of a complex regulatory system that orchestrates the state of assembly of the TJ multiprotein network. Many pathogenic bacteria exploit this system to accomplish their pathogenic strategies by ultimately modulating intestinal permeability.
The effect of delta G on the transport and oral absorption of macromolecules.
Delta G (DeltaG) is the biologically active fragment of Zonula Occludens Toxin (Zot), an absorption enhancer, that reversibly opens the tight junctions of epithelial and endothelial cells in the small intestine and brain. This study evaluates the possible use of DeltaG in enhancing the oral bioavailability of macromolecules using large paracellular markers as model agents. The transport of [(14)C]Inulin and [(14)C]PEG4000 was evaluated across Caco-2 cells with DeltaG (0, 100, 180 microg/ml). The apparent permeability coefficients (P(app)) were calculated. The in vitro toxicity of DeltaG (180 microg/ml) was assessed. Sprague Dawley rats were dosed intraduodenally (ID) with the following treatments: [(14)C]Inulin or [(14)C]PEG4000 (30 microci/kg) w/o DeltaG (720 microg/kg)/protease inhibitors (PI). Blood was collected and plasma was analyzed for radioactivity. DeltaG (180 microg/ml) increased [(14)C]Inulin and [(14)C]PEG4000 P(app) by 82.6 and 24.4%, respectively, without any toxicity. After ID administration with DeltaG/PI, C(max) and AUC were significantly (p < 0.05) increased for both Inulin and PEG4000. However, Inulin displayed greater enhancement ratios in vitro and in vivo. This study suggests that DeltaG may be used to enhance the oral bioavailability of macromolecules (e.g., proteins) after coadministration through modulation of paracellular transport.
Relationship of HLA-DQ8 and severity of celiac disease: comparison of New York and Parisian cohorts.
BACKGROUND & AIMS: Celiac disease is a polygenic disorder associated with HLA-DQ2 or HLA-DQ8, which are present in greater than 90% of patients. The disease is considered milder in the United States compared with Europe. We assessed whether differences in the frequency of HLA type may account for differences in severity of the disease by using cohorts of patients from New York and Paris. METHODS: HLA-DQ typing was performed on patients with celiac disease in New York and Paris. Clinical and pathologic data were compared between the New York and Parisian cohorts and also correlated with the different HLA types (HLA-DQ2, HLA-DQ2/-DQ8, HLA-DQ8). RESULTS: Among these patients, the disease was milder in the New York cohort compared with the Parisian cohort. There were fewer patients with a classical presentation (45% and 89%, respectively; P < 0.001) and less severe pathology (total villous atrophy, 64% and 89%, respectively; P < 0.05), and less marked intraepithelial lymphocytosis (intraepithelial leukocytes [IELs]/100 enterocytes, 48.1 and 82.5, respectively; P < 0.0001). HLA-DQ2 homozygotes were less prevalent in the New York cohort compared with the Parisian cohort (59% and 79%, respectively; P = 0.08). HLA-DQ8 alleles were more prevalent in the New York cohort compared with the Parisian cohort (41% and 21%, respectively; P = 0.026). There was, however, no difference in the clinical or pathologic parameters of severity when we compared the groups based on HLA type. CONCLUSIONS: HLA-DQ8 alleles were increased in the New York cohort of patients with celiac disease; however, this did not account for less severe manifestations of the disease.
Intestinal infections and environmental enteropathy: Working Group report of the second World Congress of Pediatric Gastroenterology, Hepatology, and Nutrition.
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Celiac disease as a cause of growth retardation in childhood.
PURPOSE OF REVIEW: Celiac disease is a syndrome characterized by damage of the small intestinal mucosa caused by the gluten fraction of wheat proteins and similar alcohol-soluble proteins (prolamines) of barley and rye in genetically susceptible subjects. The presence of gluten in these subjects leads to a self- perpetuating mucosal damage, and the elimination of gluten results in full mucosal recovery. The clinical manifestations of celiac disease are protean in nature and vary markedly with the age of the patient, the duration and extent of disease, and the presence of extraintestinal pathologic changes. In addition to the classic gastrointestinal form, a variety of other clinical manifestations of the disease have been described, including atypical and asymptomatic forms. Although the typical form of celiac disease, characterized by failure to thrive, is still the most frequent presentation in the pediatric age group, severe growth delay is less commonly seen in developed countries. RECENT FINDINGS: Recent epidemiologic studies suggest that celiac disease-associated growth retardation is becoming a tangible health problem in developing countries, where the problem has been historically overlooked. Given the protean nature of the clinical presentation of celiac disease, the diagnosis is extremely challenging and relies on a sensitive and specific algorithm that allows the identification of different manifestations of the disease. Serologic tests developed in the past decade provide a noninvasive tool for screening individuals at risk for the disease as well as the general population. SUMMARY: The current gold standard for the diagnosis of celiac disease remains histologic confirmation of the intestinal damage in serologically positive individuals. The keystone treatment of celiac disease patients is a lifelong elimination diet in which food products containing gluten are avoided.
Celiac disease--how to handle a clinical chameleon.
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Prevalence of celiac disease in at-risk and not-at-risk groups in the United States: a large multicenter study.
BACKGROUND: Celiac disease (CD) is an immune-mediated enteropathic condition triggered in genetically susceptible individuals by the ingestion of gluten. Although common in Europe, CD is thought to be rare in the United States, where there are no large epidemiologic studies of its prevalence. The aim of this study was to determine the prevalence of CD in at-risk and not-at-risk groups in the United States. METHODS: Serum antigliadin antibodies and anti-endomysial antibodies (EMA) were measured. In EMA-positive subjects, human tissue transglutaminase IgA antibodies and CD-associated human leukocyte antigen DQ2/DQ8 haplotypes were determined. Intestinal biopsy was recommended and performed whenever possible for all EMA-positive subjects. A total of 13 145 subjects were screened: 4508 first-degree and 1275 second-degree relatives of patients with biopsy-proven CD, 3236 symptomatic patients (with either gastrointestinal symptoms or a disorder associated with CD), and 4126 not-at-risk individuals. RESULTS: In at-risk groups, the prevalence of CD was 1:22 in first-degree relatives, 1:39 in second-degree relatives, and 1:56 in symptomatic patients. The overall prevalence of CD in not-at-risk groups was 1:133. All the EMA-positive subjects who underwent intestinal biopsy had lesions consistent with CD. CONCLUSIONS: Our results suggest that CD occurs frequently not only in patients with gastrointestinal symptoms, but also in first- and second-degree relatives and patients with numerous common disorders even in the absence of gastrointestinal symptoms. The prevalence of CD in symptomatic patients and not-at-risk subjects was similar to that reported in Europe. Celiac disease appears to be a more common but neglected disorder than has generally been recognized in the United States.
Effect of the biologically active fragment of zonula occludens toxin, delta G, on the intestinal paracellular transport and oral absorption of mannitol.
OBJECTIVE: Many therapeutically active agents experience low bioavailability upon oral administration due to low permeability, low solubility, interaction with efflux transporters or first pass metabolism. In general, absorption enhancers are agents that can modulate the paracellular permeability of drugs, thus, potentially increasing oral bioavailability. The objective of this study was to examine the effect of the active fragment of Zonula occludens toxin (Zot), deltaG, on the transport of a paracellular marker, mannitol, using in vitro (Caco-2 cell monolayers) and in vivo (intraduodenal administration in rats) experimental methods. METHODS: The transport of [14C]mannitol with deltaG (0, 50, 80, or 100 microg/ml) was determined across Caco-2 cells. Male Sprague-Dawley rats were assigned to receive one of the following treatments: [14C] mannitol (40 microCi/kg), [14C]mannitol/deltaG (417 microg/kg), or [14C] mannitol/deltaG/Protease inhibitors (PI). RESULTS: The mean (+/-S.E.M.) apparent mannitol permeability coefficients (P(app)) observed after incubation with 0, 50, 80, and 100 microg/ml deltaG were 3.5 (+/-0.4), 4.17 (+/-0.27), 4.33 (+/-0.61), and 9.94 (+/-0.24)x10(-6) cm/s. After oral administration, C(max) (3.8 x 10(-4) vs. 4.4 x 10(-4) mM) and AUC(0-6 h) (0.096 vs. 0.088 mM min), obtained for [14C]mannitol and [14C]mannitol/deltaG, respectively, were not statistically different. However, both C(max) (7.6 x 10(-4) mM) and AUC(0-6 h) (0.25 mM min) were significantly higher for the [14C]mannitol/deltaG/PI treatment. CONCLUSIONS: The 12 kDa fragment of Zot, deltaG, enhanced the in vitro transport and oral absorption of the paracellular marker, mannitol, in the presence of protease inhibitors (PI).
Zonula occludens toxin increases the permeability of molecular weight markers and chemotherapeutic agents across the bovine brain microvessel endothelial cells.
The purpose of this study was to examine the ability of Zonula occludens toxin (Zot) to reversibly open tight junctions in bovine brain microvessel endothelial cells (BBMECs) to enhance drug delivery via the paracellular pathway. Transport across BBMEC monolayers was examined for molecular weight markers and chemotherapeutic agents ([(14)C]sucrose, [(14)C]inulin, [(3)H]propranolol, [(3)H]doxorubicin, and [(14)C]paclitaxel) with Zot (0.0-4.0 microg/mL). TEER of monolayers was measured to assess effect and reversibility of Zot. Cell viability of BBMEC in the presence of Zot was assessed by trypan blue exclusion staining. Apparent permeability (P(app)), enhancement ratio (R), and percent increase in transport determined were statistically compared by ANOVA. A significant increase (p < 0.05) in P(app) was observed for the transport of [(14)C]sucrose, [(14)C]inulin, [(3)H]doxorubicin, and [(14)C]paclitaxel at a 4.0 microg/mL concentration of Zot. A significant concentration-dependent decrease in TEER was observed on treatment with Zot with rapid reversal to baseline after removal. Zot (4 micro/ml) was found to be nontoxic to the BBMECs after 2 hours incubation. In conclusion, Zot increased paracellular transport across the BBMEC in a reversible, concentration-dependent manner. Modulation of paracellular transport with Zot may be used to increase the brain permeability of potent central nervous system-active drugs, including anticancer agents.
Bioterrorism and biological warfare: not only a respiratory affair.
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Zonula occludens toxin acts as an adjuvant through different mucosal routes and induces protective immune responses.
Zonula occludens toxin (Zot) is produced by Vibrio cholerae and has the ability to increase mucosal permeability by reversibly affecting the structure of tight junctions. Because of this property, Zot is a promising tool for mucosal drug and antigen (Ag) delivery. Here we show that Zot acts as a mucosal adjuvant to induce long-lasting and protective immune responses upon mucosal immunization of mice. Indeed, the intranasal delivery of ovalbumin with two different recombinant forms of Zot in BALB/c mice resulted in high Ag-specific serum immunoglobulin G titers that were maintained over the course of a year. Moreover, His-Zot induced humoral and cell-mediated responses to tetanus toxoid in C57BL/6 mice and protected the mice against a systemic challenge with tetanus toxin. In addition, we found that Zot also acts as an adjuvant through the intrarectal route and that it has very low immunogenicity compared to the adjuvant Escherichia coli heat-labile enterotoxin. Finally, by using an octapeptide representing the putative binding site of Zot and of its endogenous analogue zonulin, we provide evidence that Zot may bind a mucosal receptor on nasal mucosa and may mimic an endogenous regulator of tight junctions to deliver Ags in the submucosa. In conclusion, Zot is a novel and effective mucosal adjuvant that may be useful for the development of mucosal vaccines.
Zonula occludens toxin (Zot) interferes with the induction of nasal tolerance to gliadin.
Both nasal and oral administration of soluble protein antigens (Ags) induce tolerance, a phenomenon that has hampered mucosal vaccine design. To produce active immunity the use of adjuvants co-administered with soluble Ags is required. Cholera toxin (CT) and Escherichia coli heat-labile enterotoxin (LT) were found to be powerful mucosal adjuvants, but they are not suitable for clinical use because of their associated toxicity. Therefore, there is the need to develop alternative strategies to deliver Ag in order to induce immunoprotection. Among these innovative tools, a new toxin, Zonula occludens toxin (Zot), produced by phages in toxigenic strains of Vibrio cholerae, has been recently exploited for its adjuvant activity at the mucosal level. The present study was undertaken to further highlight the adjuvant properties of Zot. The ability of Zot to induce a mucosal response to gliadin was demonstrated per serum antibody production. In our established model of systemic tolerance to gliadin, induced by its nasal administration, we found a reduced production of interferon-gamma (IFN-gamma) and interleukin-2 (IL-2) upon administration of gliadin alone. This immune suppression was reverted in mice receiving gliadin together with Zot. As previously shown, the down-regulation of Th1-like cytokines was found to be associated to a suppression of the T-cell proliferation, while such a suppression was completely reverted by Zot co-administration. In conclusion, these data confirm Zot as a good mucosal adjuvant, considering its ability to interfere with the suppression of specific cell mediated immunity, probably as a result of the increased dose and/or altered processing of Ag at mucosal level.