Effect of structural and conformation modifications, including backbone cyclization, of hydrophilic hexapeptides on their intestinal permeability and enzymatic stability

A library of 18 hexapeptide analogs was synthesized, including sub-libraries of N- or C-methylation of the parent hexapeptide Phe-Gly-Gly-Gly-Gly-Phe, as well as backbone cyclized analogs of each linear analog with various ring sizes. N- or C-methylation as well as cyclization (but not backbone cyclization) have been suggested to improve intestinal permeability and metabolic stability of peptides in general. Here we aimed to assess their applicability to hydrophilic peptides. The intestinal permeability (Papp) of the 18-peptide library was in the range of 0.2-6.8 x 10-6 cm/sec. Based on several tests, we concluded that the absorption mechanism of all tested analogs is paracellular, regardless of the structural or conformational modifications. In all cases, backbone cyclization increased Papp (5-fold) in comparison to the linear analogs due to the smaller 3D size and also dramatically decreased peptide proteolysis by brush border enzymes. N- or C-methylation did not enhance the permeability of the linear analogs in this series.     

Transcutaneous immunization with a synthetic hexasaccharide-protein conjugate induces anti-Vibrio cholerae lipopolysaccharide responses in mice

Antibodies specific for Vibrio cholerae lipopolysaccaride (LPS) are common in humans recovering from cholera, and constitute a primary component of the vibriocidal response, a serum complement-mediated bacteriocidal response correlated with protection against cholera. In order to determine whether transcutaneous immunization (TCI) with a V. cholerae neoglycoconjugate (CHO-BSA) comprised of a synthetic terminal hexasaccharide of the O-specific polysaccharide of V. cholerae O1 (Ogawa) conjugated with bovine serum albumin (BSA) could induce anti-V. cholerae LPS and vibriocidal responses, we applied CHO-BSA transcutaneously in the presence or absence of the immune adjuvant cholera toxin (CT) to mice. Transcutaneously applied neoglycoconjugate elicited prominent V. cholerae specific LPS IgG responses in the presence of CT, but not IgM or IgA responses. CT applied on the skin induced strong IgG and IgA serum responses. TCI with neoglycoconjugate did not elicit detectable vibriocidal responses, protection in a mouse challenge assay, or stool anti-V. cholerae IgA responses, irrespective of the presence or absence of CT. Our results suggest that transcutaneously applied synthetic V. cholerae neoglycoconjugate is safe and immunogenic, but predominantly induces systemic LPS responses of the IgG isotype.     

Enhanced immunogenicity of an oral inactivated cholera vaccine in infants in Bangladesh obtained by zinc supplementation and by temporary withholding breast-feeding

The killed oral cholera vaccine Dukoral is recommended for adults and only children over 2 years of age, although cholera is seen frequently in younger children and there is an urgent need for a vaccine for them.     

Distribution of the Escherichia coli Common Pilus among Diverse Strains of Human Enterotoxigenic E. coli

The Escherichia coli common pilus (ECP) is produced by commensal and pathogenic E. coli strains. This pilus is unrelated to any of the known colonization factors (CFs) of enterotoxigenic E. coli (ETEC). In this study, we investigated the distribution and production of ECP among a collection of 136 human CF-positive and CF-negative ETEC strains of different geographic origins. The major pilus subunit gene, ecpA, was found in 109 (80%) of these strains, suggesting that it is widely distributed among ETEC strains. Phenotypic analysis of a subset of 43 strains chosen randomly showed that 58% of them produced ECP independently of the presence or absence of CFs, a percentage even higher than that of the most prevalent CFs. These data suggest an important role for ECP in the biology of ETEC, particularly in CF-negative strains, and in human infection.Enterotoxigenic Escherichia coli (ETEC) is an important cause of diarrheal disease and mortality for children living in developing countries (11). The presence of ETEC in these areas is associated with a lack of sanitation or poor sanitation and the consumption of contaminated water or food. The main virulence factors of ETEC are a heat-labile (LT) and/or a heat-stable (ST) enterotoxin and multiple adhesive pili called colonization factors (CFs) (1, 7), which are produced in the small intestine and can cause life-threatening, cholera-like watery diarrhea (7). Since the early 1970s, more than 25 different CFs have been reported in ETEC strains of diverse geographic origins, and the prevalence of these pili differs by geographic region (7, 11). Studies of the prevalence and distribution of CFs among ETEC strains worldwide have shown that the most common CFs are CFA/I and combinations of E. coli surface antigens CS1, CS2, and CS3 or of antigens CS4, CS5, and CS6. Approximately 50% of ETEC strains contain at least one of these CFs (7), leaving 50% of strains that do not produce any of the CFs known or characterized so far. The presence of type IV pili, which are associated with host colonization and virulence in many gram-negative bacteria, has also been demonstrated in a significant number (30 to 50%, depending on the geographic source) of ETEC strains, including strains that do not harbor any of the known CFs. These pili provide a mechanism for the organisms to colonize the human gut and establish gastrointestinal disease. Epidemiological studies have shown that protective immunity, attributed to the antigenic variety of the CFs produced, can be achieved through multiple infections. Thus, it is believed that vaccines aimed at preventing ETEC infections, particularly in the young population and travelers, should contain the immunogenic B subunit of the LT and a combination of the most common CFs (7, 9, 10).Previously, it was reported that meningitis-associated E. coli strains, and not other E. coli pathogroups, were able to assemble a “meningitis-associated temperature-dependent pilus” (Mat) after growth at 20°C in Luria-Bertani (LB) medium. The major pilus subunit of the Mat pilus is encoded by the yagZ gene, commonly found in all E. coli strains. Recently, our laboratory reported that most (75%) strains of human and animal E. coli pathogroups (including ETEC), as well as commensal E. coli strains, produce at 37°C a pilus adhesive structure composed of a major 21-kDa protein pilin subunit corresponding to the product of the yagZ gene (8). Because this gene was demonstrated to be widely distributed and highly conserved among E. coli strains, and because production of the pili was shown in the major E. coli pathovars, it was proposed that the pilus be renamed “E. coli common pilus,” or ECP, and that the gene encoding the pilin subunit be designated ecpA. A role for ECP in adherence to cultured human epithelial cells was demonstrated in enterohemorrhagic E. coli (EHEC) O157:H7 and commensal E. coli strains (8).ECP is not related to any of the known ETEC CFs. The present study was carried out to further investigate the presence of ecpA and to determine the production of ECP in a collection of human ETEC strains that had previously been characterized as CF positive or CF negative. We found ECP production in both groups of strains at rates comparable to those found for the most common CFs. Our data suggest that the production of ECP in ETEC strains may contribute to the adhesive properties of this organism and may represent a target for vaccine development and the prevention of ETEC infections.     

Postgenomic approaches to cholera vaccine development

Cholera remains an important public health threat. A cholera vaccine that provides durable protection at the mucosal surface, especially among children in endemic settings, is urgently needed. The availability of the complete genome sequence of a clinical isolate of Vibrio cholerae O1 El Tor has allowed for comparative and functional genomic approaches in the study of cholera. This work holds promise for the identification of bacterial targets of protective human immune responses and may contribute to the development of a new generation of cholera vaccines.     

A case of extranasopharyngeal angiofibroma of the ethmoid sinus: a distinct clinical entity at an unusual site

Angiofibromas of the head and neck usually arise from the nasopharynx in adolescent boys. The term extranasopharyngeal angiofibroma (ENA) has been applied to vascular fibrous nodules that arise outside the nasopharynx. Because the clinical characteristics of ENA are not consistent with those of juvenile nasopharyngeal angiofibroma (JNA), diagnosis of the former can be challenging. Biopsy is ill advised in a patient with an ENA because it might result in brisk bleeding. A high index of suspicion and a methodical evaluation are essential in establishing the proper diagnosis and treatment. We report the rare case of a 16-year-old girl who presented with a pinkish lobulated mass in her left nostril that had arisen from the anterior ethmoid sinus. The mass was removed via a lateral rhinotomy approach. Postoperative histopathologic analysis identified it as an angiofibroma. To best of our knowledge, only 9 cases of ENA arising from the ethmoid sinus have been previously reported in the English-language literature.     

5-flourouracil cardiotoxicity - an elusive cardiopathy: case report

5-Flourouracil (5-FU) is an S-phase specific, synthetic pyrimidine antimetabolite. It is a frequently administered chemotherapeutic agent for a variety of malignant lesions, either singly or in multidrug regimens. Its adverse side effects involving bone marrow, skin, mucous membranes, GIT and CNS are well known, whereas its cardiotoxicity is relatively uncommon and occurs in 1.2-18%. Keywords: 5-Flourouracil, Cardiotoxicity, S-phase specific.     

Growth retardation and altered autonomic control in mice lacking brain serotonin

Serotonin synthesis in mammals is initiated by 2 distinct tryptophan hydroxylases (TPH), TPH1 and TPH2. By genetically ablating TPH2, we created mice (Tph2^−/−) that lack serotonin in the central nervous system. Surprisingly, these mice can be born and survive until adulthood. However, depletion of serotonin signaling in the brain leads to growth retardation and 50% lethality in the first 4 weeks of postnatal life. Telemetric monitoring revealed more extended daytime sleep, suppressed respiration, altered body temperature control, and decreased blood pressure (BP) and heart rate (HR) during nighttime in Tph2^−/− mice. Moreover, Tph2^−/− females, despite being fertile and producing milk, exhibit impaired maternal care leading to poor survival of their pups. These data confirm that the majority of central serotonin is generated by TPH2. TPH2-derived serotonin is involved in the regulation of behavior and autonomic pathways but is not essential for adult life.