The receptor binding domain of apolipoprotein E, linked to a model class A amphipathic helix, enhances internalization and degradation of LDL by fibroblasts.

Human apolipoprotein E (apo E) consists of two distinct domains, the lipid-associating domain (residues 192-299) and the globular domain (residues 1-191) which contains the LDL receptor (LDLR) binding site (residues 129-169). To test the hypothesis that an arginine-rich apo E receptor binding domain (residues 141-150) is sufficient to enhance low-density lipoprotein (LDL) uptake and clearance when covalently linked to a class A amphipathic helix, a peptide in which the receptor binding domain of human apo E, LRKLRKRLLR (hApoE[141-150]), is linked to 18A, a well-characterized high-affinity lipid-associating peptide (DWLKAFYDKVAEKLKEAF), we synthesized the peptide hApoE[141-150]-18A (hE18A) and its end-protected analogue, Ac-hE18A-NH(2). The importance of positively charged residues and the role of the hydrophobic residues in the receptor binding domain were also studied using four analogues.

Imaging and tissue biodistribution of 99mTc-labeled adenovirus knob (serotype 5).

Hepatic sequestration of systemically administered adenoviral vectors reduces the number of viral particles available for delivery to other tissues. The biological basis of this phenomenon was investigated using a new in vivo technique which permitted imaging in real time. Recombinant adenovirus serotype 5 knob (Ad5K) was radiolabeled with the gamma-emitter 99mTc (half-life = 6 h).

The lipid-free structure of apolipoprotein A-I: effects of amino-terminal deletions.

Deletion mutants of human apolipoprotein A-I (apo hA-I) have been produced from a bacterial expression system to explore the function of the specific domains comprising residues 1-43, 1-65, 88-98, and 187-243, respectively, in the lipid-free conformation and in the lipid-binding mechanism of apo hA-I. Initial studies on apo Delta(1-43)A-I and apo Delta(187-243)A-I have already been reported. To aid purification of these mutants, a histidine-containing N-terminal extension was incorporated (+his); in cases where comparison with the (-his) construct was possible, little effect on the physical properties due to the (+his) extension was found.

Structural analysis of apolipoprotein A-I: effects of amino- and carboxy-terminal deletions on the lipid-free structure.

An amino-terminal deletion mutant (residues 1-43) and a carboxy-terminal deletion mutant (residues 187-243) of human apoliprotein A-I (apo hA-I) have been produced from a bacterial expression system to explore the importance of the missing residues for the conformation of apo hA-I. Our focus has been to study the lipid-free structure of apo hA-I to understand how discrete domains influence the conformational plasticity of the protein and, by inference, the mechanism of lipid binding. All spectral and physical measurements indicate that both apo delta(1-43)A-I and apo delta(187-243)A-I have folded, tertiary structures.

Interleukin-12 alters helper T-cell subsets and antibody profiles induced by the mucosal adjuvant cholera toxin.

We have shown that systemic administration of rmIL-12 could trigger Th1-type responses to a protein antigen delivered orally with CT as mucosal adjuvant. The most striking finding was that IL-12 could retain its regulatory effects when orally administered and could redirect the immune response to the oral vaccine toward a Th1-type. However, regulation by orally administered IL-12 differed from parenteral treatment with IL-12 since only the latter treatment affected mucosal S-IgA responses.

Hapten-induced model of murine inflammatory bowel disease: mucosa immune responses and protection by tolerance.

We report here a murine model for experimental chronic colitis where administration of trinitrobenzene sulfonic acid (TNBS) in 50% ethanol induced inflammation of large intestine in susceptible (C3H/HeJ and BALB/c) but not resistant (C57BL/6 and DBA/2) mouse strains. We queried whether mucosal trinitrophenyl (TNP)-specific B cell responses were induced in mice with TNBS-induced colitis, and if induction of tolerance to TNBS by oral administration of this hapten protected mice from development of colitis. Isotypes and subclasses of polyclonal and TNP-specific Ab-forming cells (AFC) were assessed in mucosal and peripheral lymphoid tissues of C3H/HeJ mice with TNBS-induced colitis.

Interaction of model class A1, class A2, and class Y amphipathic helical peptides with membranes.

To test the hypothesis that differences in the lipid affinity of exchangeable apolipoproteins are due to the presence of different classes of amphipathic alpha-helical motifs which differ primarily in the distribution of charged amino acid residues, we designed and synthesized model peptides mimicking class A1, class A2, and class Y amphipathic helices present in these apolipoproteins. Both class A1 and class A2 helices have positive residues at the polar-nonpolar interface and negative residues at the center of the polar face. However, clustering of positive and negative residues is less exact in class A1 compared to class A2 helices.

Oral immunization of interleukin-4 (IL-4) knockout mice with a recombinant Salmonella strain or cholera toxin reveals that CD4+ Th2 cells producing IL-6 and IL-10 are associated with mucosal immunoglobulin A responses.

Mucosal immunoglobulin A (IgA) responses are often associated with Th2-type cells and derived cytokines, and interleukin-4 (IL-4) knockout (IL-4-/-) mice with impaired Th2 cells respond poorly to oral antigens. However, we have noted that IL-4-/- mice have normal mucosal IgA levels, which led us to query whether different oral delivery systems could elicit mucosal immunity. Two oral regimens were used: (i) a live recombinant Salmonella strain which expresses fragment C (ToxC) of tetanus toxin, and (ii) soluble tetanus toxoid (TT) with cholera toxin (CT) as an adjuvant.

Absence of exogenous interleukin-4-induced apoptosis of gingival macrophages may contribute to chronic inflammation in periodontal diseases.

Inflamed gingival tissues are enriched in macrophages (MOs) and CD4-positive T cells; however, T helper-type cytokines such as interleukin (IL)-2 and IL-4 are absent. Therefore, we investigated whether a relationship exists between IL-4 receptor (IL-4R) expression and MO persistence in the absence of exogenous IL-4. Gingival MOs, when compared with monocyte(MN)/MOs from peripheral blood mononuclear cells, expressed high levels of IL-4R mRNA.

Polarized Th2 cytokine expression by both mucosal gamma delta and alpha beta T cells.

Currently only limited information is available as to why dominant IgA isotype responses are supported by mucosal T cells in effector tissues. To address this issue directly, gamma delta and alpha beta T cells were isolated from the submandibular gland (SMG) of mice as an example of mucosal effector tissues. Freshly isolated CD3+ T cells from this tissue contained relatively high numbers of activated cells [approximately 10% interleukin-2 receptor (IL-2R)+ cells and 15% of cells in cycle stages S and G2 + M], of which 25% and 75% were gamma delta and alpha beta T cells, respectively.

Ventricular to pulmonary artery connections: generalizations.

The basic information about surgically created ventricular to pulmonary artery connections is condensed into five general headings. The information is useful, but is no substitute for the technically competent, well-informed, thoughtful, and intuitive cardiac surgeon.

Molecular basis for prokaryotic specificity of magainin-induced lysis.

Magainins and mastoparans are examples of peptide antibiotics and peptide venoms, respectively. They have been grouped together as class L amphipathic helixes [Segrest, J.P.

Case report: milk-alkali syndrome and pancreatitis.

The relation between hypercalcemia and pancreatitis, first described in patients with hyperparathyroidism, is controversial. Other causes of hypercalcemia also have been associated with pancreatitis. In this report, the authors describe a patient with pancreatitis and the milk-alkali syndrome who had the classic triad of hypercalcemia, alkalosis, and renal insufficiency.

Oral vaccine models: multiple delivery systems employing tetanus toxoid.

We have not yet directly examined the Th cell responses induced by using Salmonella/BRD 847 as a vector nor have we performed these experiments following immunization with microspheres. However, production of high serum levels of antigen-specific IgG1 may be indicative of a Th2-type response, whereas high serum levels of IgG2a may reflect a Th1-type response. An important issue in using various oral delivery systems is whether the system(s) employed affects the Th cell response to the same antigen.

The enteric nervous and immune systems: interactions for mucosal immunity and inflammation.

A number of anatomical studies have demonstrated the presence of peptidergic nerve fibers infiltrating mucosal lymphoid tissues. The exact mechanisms of how neuropeptides are released to affect these lymphoid sites are unclear, but radiolabeled binding studies have shown that mucosal leukocytes bear a number of neuropeptide receptors on their cell surfaces capable of responding to neural signals. The presence of neuropeptide-containing fibers and the ability to receive neural signals suggest that mucosal lymphocytes can be influenced by neurogenic mediators.

Interactions of synthetic peptide analogs of the class A amphipathic helix with lipids. Evidence for the snorkel hypothesis.

Class A amphipathic helixes present in exchangeable plasma apolipoproteins are characterized by the location of positively charged amino acid residues at the non-polar-polar interface and negatively charged amino acid residues at the center of the polar face. The objectives of the present study were: (i) to investigate the role of hydrocarbon side chain length of the interfacial positively charged amino acid residues in the lipid affinity of class A amphipathic helixes, and (ii) to investigate the importance of the nature of interfacial charge in the lipid affinity of class A amphipathic helixes. Toward this end, lipid interactions of the following two analogs of the class A amphipathic helix, Ac-18A-NH2 (acetyl-Asp-Trp-Leu-Lys-Ala-Phe-Tyr- Asp-Lys-Val-Ala-Glu-Lys-Leu-Lys-Glu-Ala-Phe-NH2), and Ac-18A(Lys > Haa)-NH2 (acetyl-Asp-Trp-Leu-Haa-Ala-Phe-Tyr-Asp-Haa-Val-Ala-Glu-Haa-Leu-Haa-Glu- Ala-Phe-NH2) (Haa = homoaminoalanine), were studied.

alpha beta T cell receptor-positive intraepithelial lymphocytes with CD4+, CD8- and CD4+, CD8+ phenotypes from orally immunized mice provide Th2-like function for B cell responses.

Our previous studies have shown that murine alpha beta TCR+ intraepithelial lymphocytes (IEL) contained T cells that can provide B cell help. In this study, we have examined the three subsets of alpha beta TCR+ IEL for Ag-specific helper function and cytokine production because alpha beta TCR+ IEL are divisible into three subsets including CD4+, CD8- T cells, CD4+, CD8+ double positive (DP) T cells, and CD4-, CD8+ T cells. When these three subsets of alpha beta TCR+ IEL from C3H/HeN mice (H-2k) orally immunized with SRBC were cultured with splenic B cells, adherent cells, and SRBC, both CD4+, CD8- and DP T cell fractions supported IgM, IgG1, and IgA anti-SRBC responses, whereas the CD4-, CD8+ T cell subset did not exhibit helper function.

Enhancing effect of cholera toxin on interleukin-6 secretion by IEC-6 intestinal epithelial cells: mode of action and augmenting effect of inflammatory cytokines.

Oral administration of cholera toxin (CT) induces a strong mucosal immune response to CT as well as having a potent adjuvant effect. Since one of the first cell types to encounter CT during cholera infection or after oral administration is the epithelial cell, we studied the effect of CT on interleukin-6 (IL-6) secretion by the rat intestinal epithelial cell line IEC-6. CT was found to rapidly enhance IL-6 secretion and IL-6 gene expression by these cells.

Optimizing oral vaccines: induction of systemic and mucosal B-cell and antibody responses to tetanus toxoid by use of cholera toxin as an adjuvant.

Cholera toxin (CT) is an effective mucosal antigen and acts as an adjuvant when given orally with various antigens; however, few studies have compared the levels of antibody responses to CT and coadministered protein in systemic and mucosal tissues. In this study, we used tetanus toxoid (TT) for assessment of immune responses. Time course and dose-response studies established that 250 micrograms of TT given orally with 10 micrograms of CT three times at weekly intervals induced high serum and gastrointestinal tract anti-TT and anti-CT antibody responses.

The future of cardiac surgery.

Cardiac surgery in the future will probably be different than it is today. It is unlikely that the general changes that have occurred in the demands on cardiac surgeons and cardiologists and their responses to them will be reversed. Therefore, it is likely that the role of the primary care physician in decision-making will increase.

When and how to include arch repair in patients with acute dissections involving the ascending aorta.

The arch should be included in the repair in patients with acute aortic dissection involving the ascending aorta when (1) the intimal tear is in the arch, (2) rupture of the arch has occurred, (3) the outer wall of the false channel in the arch is tenuous, and (4) the inner wall of the false channel is fragmented. A technique has evolved for patients with acute aortic dissection involving the arch that seems to minimize the difficulties and risks of including the arch, when indicated. Its place will best be determined by additional, prospective, and preferably multi-institutional testing.

Clinical outcomes after the arterial switch operation for transposition. Patient, support, procedural, and institutional risk factors. Congenital Heart Surgeons Society.

Background: As the probability increases that the arterial switch operation is optimal treatment for transposition, detailed information about outcomes and the circumstances in which they are suboptimal becomes important.

Methods And Results: A multi-institutional prospective study with annual detailed follow-up included 513 neonates with simple transposition or transposition and ventricular septal defect entering for diagnosis and treatment at < 15 days of age and undergoing an arterial switch repair. The 1-month and 1- and 5-year survivals were 84%, 82%, and 82%, respectively.

Combined femoral pseudoaneurysm and arteriovenous fistula: diagnosis by Doppler color flow mapping.

This case report is the first to describe a combined femoral pseudoaneurysm and arteriovenous fistula resulting from a cardiac catheterization, diagnosed by color Doppler.