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Biomedical subjects

M Fridkin

Publications and source records attributed to M Fridkin.

At least 73 records · Page 4Linked to original sources
PubMed1998

Multiple actions of a hybrid PACAP antagonist: neuronal cell killing and inhibition of sperm motility.

Pituitary stimulating adenylate cyclase (PACAP) is a major regulatory peptide with two active molecular forms: PACAP-27 and PACAP-38. Both molecular forms promote neuronal survival and protect against neurotoxicity. Based on our previous hybrid peptide strategy in designing vasoactive intestinal peptide (VIP) antagonists, novel PACAP analogues were synthesized (neurotensin6-11 PACAP7-27 and neurotensin6-11 PACAP7-38). In addition to the hybrid modification, the methionine in position 17 was replaced by norleucine (Nle). Treatment of rat cerebral cortical cultures for five days with the putative PACAP antagonists (1 nM) resulted in a 35-45% reduction in neuronal cell counts as compared to controls. Neuronal cell death was already obtained at picomolar concentrations for the neurotensin6-11 PACAP7-27 antagonist with 70% death at 10(-8) M. Co-administration of the PACAP hybrid analogue with picomolar amounts of PACAP-27 or Nle17-PACAP-27 attenuated the reduction in neuronal cell counts. While the protective effects of both analogues exhibited a peak at 1 pM concentrations, the Nle-containing agonist displayed a broader range of active concentrations (10(-12)M-10(-9) M). The putative PACAP antagonist also inhibited sperm motility (golden hamster) in a dose-dependent manner as assessed in vitro. Complete inhibition was observed at 10 microM, suggesting a role for PACAP in sperm motility and sexual function. Thus, previous findings of a large number of PACAP and PACAP receptors in the nervous system and the reproductive system are now correlated with a function in neuronal survival and sperm motility. The structure-activity studies suggest that the methionine in position 17 and the first six amino acids are important in the determination of PACAP activity, knowledge that may facilitate PACAP-based drug design.

Amino Acid Sequence↗
PubMed1998

Vasoactive intestinal peptide and pituitary adenylate cyclase-activating polypeptide inhibit tumor necrosis factor alpha transcriptional activation by regulating nuclear factor-kB and cAMP response element-binding protein/c-Jun.

Tumor necrosis factor alpha (TNFalpha), an early cytokine produced by activated macrophages, plays an essential role in normal and pathological inflammatory reactions. The excessive production of TNFalpha is prevented by the so-called "macrophage-deactivating factors." This study examines the role of two structurally related neuropeptides, the vasoactive intestinal peptide (VIP) and the pituitary adenylate cyclase-activating peptide (PACAP), as inhibitors of TNFalpha. Both VIP and PACAP inhibit TNFalpha production from lipopolysaccharide-stimulated RAW 246.7 cells in a dose- and time-dependent manner. Although the activated cells express mRNA for all three VIP/PACAP receptors, agonist and antagonist studies indicate that the major receptor involved is VIP1R. VIP/PACAP inhibit TNFalpha gene expression by affecting both NF-kB binding and the composition of the cAMP responsive element binding complex (CREB/c-Jun). Two transduction pathways, a cAMP-dependent and a cAMP-independent pathway, are involved in the inhibition of TNFalpha gene expression and appear to differentially regulate the transcriptional factors involved. Because TNFalpha plays a central role in various inflammatory diseases such as endotoxic shock, multiple sclerosis, cerebral malaria, and various autoimmune conditions, the down-regulatory effect of VIP/PACAP may have a significant therapeutic potential.

Animals↗
PubMed1998

IL-2 induces T cell adherence to extracellular matrix: inhibition of adherence and migration by IL-2 peptides generated by leukocyte elastase.

Migration of inflammatory cells requires cell adhesion and their subsequent detachment from the extracellular matrix (ECM). Leukocyte activation and migration must be terminated to stop inflammation. Here, we report that IL-2 enhances human T cell adherence to laminin, collagen type IV, and fibronectin (FN). In contrast, neutrophil elastase, an enzyme activated during inflammation, degrades IL-2 to yield IL-2 fractions that inhibit IL-2-induced T cell adhesion to FN. The amino acid composition of two of these IL-2 fractions, which appear to block T cell adherence to FN, were analyzed, and three peptides were consequently synthesized. The three peptides IVL, RMLT, and EFLNRWIT, but not the corresponding inversely synthesized peptides, inhibited T cell adhesion to FN induced by a variety of activators: IL-2, IL-7, macrophage inflammatory protein (MIP)-1beta, and PMA, as well as anti-CD3 and anti-beta1 integrin-activating mAb. Moreover, these IL-2 peptides inhibited T cell chemotaxis via FN-coated membranes induced by IL-2 and MIP-1beta. Inhibition of T cell adherence and migration apparently involves abrogation of the rearrangement of the T cell actin cytoskeleton. Thus, the migrating immune cells, the cytokines, and the ECM can create a functional relationship in which both inflammation-inducing signals and inhibitory molecules of immune responses can coexist; the enzymatic products of IL-2 may serve as natural feedback inhibitors of inflammation.

Actins↗
PubMed1998

Single amino acid analogs of a myasthenogenic peptide modulate specific T cell responses and prevent the induction of experimental autoimmune myasthenia gravis.

Peptide p259-271 of the human acetylcholine receptor alpha-subunit, preferentially stimulates T cells of patients with myasthenia gravis (MG) and is an immunodominant epitope for T cells of BALB/c mice. A p259-271 specific T cell line of BALB/c origin was established and was shown to induce experimental MG in naive mice. Seven analogs of p259-271 were synthesized, and two of them were found to inhibit the p259-271 specific proliferative responses of the line and of p259-271 primed lymph node cells. Moreover, the most efficient inhibitor, analog 262Lys, prevented the MG related manifestations in mice inoculated with the line, and might be of potential value for the treatment of MG.

Amino Acid Sequence↗
PubMed1998

Effect of serum amyloid A on selected in vitro functions of isolated human neutrophils.

Serum amyloid A (SAA) is an acute phase reactant whose levels in the blood rise as part of the body's response to stress and inflammation. Previous studies have suggested that SAA may carry an anti-inflammatory potential. We evaluated the effects of SAA on human neutrophils activated by N-formyl-methionyl-leucyl-phenylalanine (fMLP) in vitro. At concentrations higher than 10 microg/mL, SAA inhibited neutrophil myeloperoxidase (MPO) release. This effect was located in the N-terminal--that is, amino acid residues 1-14--of the SAA molecule. Directed neutrophil migration was inhibited at the same SAA concentrations. Several amino acid residues (1-14, 15-104, 83-104) contributed to this effect. Neutrophil O2- production was inhibited at low concentrations of SAA (0.1 to 1 microg/ml) and was stimulated at concentrations higher than 50 microg/mL. Neutrophil O2- production induced by phorbol myristate acetate (PMA) and O2- generated by the xanthine-xanthine oxidase reaction were not affected by SAA. These results add to previous data suggesting that SAA, at concentrations recorded in the serum during inflammation, modulates neutrophil function; thus it may play a role in the down-regulation of the inflammatory process.

Adult↗
PubMed1998

Protection against developmental deficiencies by a lipophilic VIP analogue.

Stearyl-Nle-VIP (SNV) is a novel agonist of vasoactive intestinal peptide (VIP) exhibiting a 100-fold greater potency than the parent molecule and specificity for a receptor associated with neuronal survival. Here, the developmental and protective effects of SNV were investigated in vivo using two models of developmental retardation, hypoxia and cholinergic blockade. In both cases chronic administration of SNV during development provided protective effects. Water maze experiments on the weaned animals have demonstrated a prophylactic action for SNV and enhancement of spatial memory in animals exposed to a cholinotoxin. SNV may act by providing neuroprotection, thereby improving cognitive functions. This work is dedicated to Prof. R.J. Wurtman whose inspiration and leadership in the field of neuroscience and cognition is beyond comparison.

Aging↗
PubMed1998

Involvement of pituitary adenylate cyclase-activating polypeptide II vasoactive intestinal peptide 2 receptor in mouse neocortical astrocytogenesis.

At the end of neuronal migration, the neopallial germinative zone produces glial cells destined to colonize the upper layers of neocortex. High densities of binding sites for vasoactive intestinal peptide (VIP) have been found in the rodent germinative zone just after completion of neuronal migration, suggesting a possible role of VIP in neocortical astrocytogenesis. In the present study, administration of a VIP antagonist at embryonic days 17 and 18 to pregnant mice was followed by a dramatic depletion of astrocytes in the upper cortical layer of the offspring. The depletion of astrocytes was dose-dependent, with a 42% reduction in the density of astrocytes observed with 50 microg of antagonist. The antagonist effect was reversed by cotreatment with VIP or pituitary adenylate cyclase-activating polypeptide (PACAP), suggesting the involvement of a receptor common to these two neuropeptides. VIP antagonist-induced inhibition of astrocytogenesis was also blocked by Ro 25-1553, a long-acting cyclic VIP analogue selective for the PACAP II VIP2 receptor subclass. Our results demonstrate that VIP and/or PACAP play a crucial physiological role in neocortical astrocytogenesis, possibly through interaction with PACAP II VIP2 receptors.

Animals↗
PubMed1998

Peptides derived from human C-reactive protein inhibit the enzymatic activities of human leukocyte elastase and cathepsin G: use of overlapping peptide sequences to identify a unique inhibitor.

Ten overlapping 15-mer peptides, spanning the entire inner disulfide loop of human C-reactive protein (residues 36-97), were used to isolate a potent inhibitor of the enzymes human leukocyte elastase and human leukocyte cathepsin G, which are associated with chronic inflammatory tissue damage. In contrast to the inability of intact C-reactive protein to inhibit both enzymes, the synthetic peptide E62ILIFWSKDIGYSFT76 inhibited leukocyte elastase (Ki = 0.18 microM) and cathepsin G (Ki = 0.25 microM) at concentrations far lower than the acute-phase concentration of C-reactive protein. Several peptide-enzyme binding motifs were elucidated by structure-function studies, with the Glu62 residue being crucial in establishing long-range subsite interactions. Peptides derived from C-reactive protein, which may be generated in vivo by neutrophil-mediated proteolysis as part of a complex regulatory homeostatic mechanism, may play an important role in regulating the activity of matrix-degrading enzymes, specifically at sites of inflammation. The present results thus may shed additional insight on the physiological functions of the major acute-phase reactant C-reactive protein, and perhaps be used as a basis for the design of novel therapeutic substances.

Amino Acid Sequence↗
PubMed1998

Activity-dependent neurotrophic factor: structure-activity relationships of femtomolar-acting peptides.

Activity-dependent neurotrophic factor (ADNF) is a glia-derived protein that is neuroprotective at femtomolar concentrations. A 14-amino acid peptide of ADNF (ADNF-14) has been reported that protects cultured neurons from multiple neurotoxins. Structure-activity relationships of peptides related to ADNF-14 now have been determined. A 9-amino acid core peptide (ADNF-9) has been identified that has greater potency and a broader effective concentration range (10(-16) to 10(-13) M) than ADNF or ADNF-14 in preventing cell death associated with tetrodotoxin treatment of cerebral cortical cultures. Deletions or conservative amino acid substitutions to ADNF-9 resulted in reduced potency, narrower effective concentration range and/or decreased efficacy. Removal of the N-terminal serine or the COOH-terminal isoleucine-proline-alanine from ADNF-9 produced a significant reduction in survival-promoting activity. Comparative studies of ADNF-9 action in mixed (glia plus neurons) vs. glia-depleted neuronal cultures indicated that ADNF-9 can act directly on neurons, although the potency of the peptide was 10,000-fold greater in mixed cultures. Kinetic studies showed that exposure to ADNF-9 for only 2 hr was sufficient to produce a 4-day protection against the cell-killing action of tetrodotoxin. Treatment with bafilomycin A1 (an inhibitor of receptor-mediated endocytosis) for 2 hr prevented the ADNF- and ADNF-9-mediated neuroprotection. ADNF-9, like ADNF-14, was neuroprotective against N-methyl-D-aspartate and the beta-amyloid peptide (amino acids 25-35), and had a much broader range of effective concentrations than ADNF-14. These studies identify ADNF-9 as an attractive lead compound for the development of therapeutic agents against neurodegenerative diseases.

Animals↗
PubMed1997

Identification of shared tumor-associated antigen peptides between two spontaneous lung carcinomas.

CTLs recognize antigenic peptides bound to MHC class I Ags on the cell surface of tumor cells. Tumor-associated Ag (TAA) peptides are 8 to 10 amino acids long and can be derived from normal, mutated, or viral proteins. The majority of T cell-defined Ags have been identified in human melanoma cells. These were shown to be commonly expressed by different allogeneic melanomas that share the same MHC molecule. We have recently isolated Kb-restricted TAA peptides, which are mutations of the gap junction protein connexin 37, from the spontaneous C57BL/6 Lewis lung carcinoma (3LL). These peptides, named MUT 1 and MUT 2, serve as CTL epitopes and can induce CTL activity in vivo. Using CTL cross-reaction assays, peptide extraction, HPLC fractionation, and reverse transcriptase-PCR amplification, we show that clones of another spontaneous C57BL/6 lung carcinoma, CMT 64, share TAA peptides with the 3LL carcinoma. Vaccination with synthetic MUT 1 or MUT 2 induces CTLs that efficiently lyse CMT 64-derived clones, protects mice from CMT 64 metastasis, and affords therapy of established CMT 64 metastases. Hence, shared CTL epitopes exist between two spontaneous murine lung carcinomas.

Animals↗
PubMed1997

Vasoactive intestinal peptide prevents excitotoxic cell death in the murine developing brain.

Excitotoxic damage may be a critical factor in the formation of brain lesions associated with cerebral palsy. When injected at birth, the glutamatergic analog ibotenate induces mouse brain lesions that strikingly mimic human microgyria. When ibotenate is injected at postnatal day 5, it produces transcortical necrosis and white matter cysts that mimic human perinatal hypoxic-like lesions. Vasoactive intestinal peptide (VIP) has potent growth-related actions and neuroprotective properties that influence mitosis and neuronal survival in culture. The goal of this study was to assess the protective role of VIP against excitotoxic lesions induced by ibotenate in developing mouse brain. VIP cotreatment reduced ibotenate-induced microgyric-like cortical lesions and white matter cysts by up to 77 and 85%, respectively. VIP protective effects were reproduced by a peptide derived from activity-dependent neurotrophic factor (ADNF), a trophic factor released by VIP-stimulated astrocytes, and by stearyl norleucine VIP, a specific VIP agonist that does not activate adenylate cyclase. Neither forskolin, an adenylate cyclase activator, nor pituitary adenylate cyclase-activating peptide, provided VIP-like protection. VIP and neurotrophic analogs, acting through a cAMP-independent mechanism and inducing ADNF release, could represent new avenues in the understanding and prevention of human cerebral palsy.

Animals↗
PubMed1997

A peptide composed of tandem analogs of two myasthenogenic T cell epitopes interferes with specific autoimmune responses.

Myasthenia gravis (MG) is a T cell-regulated, antibody-mediated autoimmune disease. Two peptides representing sequences of the human acetylcholine receptor alpha-subunit, p195-212 and p259-271, were previously shown to stimulate peripheral blood lymphocytes of patients with MG and were found to be immunodominant T cell epitopes in SJL and BALB/c mice, respectively. Single amino acid substituted analogs of p195-212 (analog Ala-207) and p259-271 (analog Lys-262) were synthesized. We showed that analogs Ala-207 and Lys-262 inhibited, in vitro and in vivo, the proliferative responses of T cell lines specific to the relevant peptide and lymph node cells of mice immunized to p195-212 and p259-271, respectively. To inhibit T cell responses to both peptides (p195-212 and p259-271), we synthesized dual analogs composed of the tandemly arranged two single (Ala-207 and Lys-262) analogs (dual analog) either sequentially (Ala-207-Lys-262) or reciprocally (Lys-262-Ala-207). In the present study, we report that both dual analogs could bind to major histocompatibility complex class II molecules on antigen-presenting cells of SJL and BALB/c mice. Analog Lys-262-Ala-207, which bound more efficiently to major histocompatibility complex class II molecules, was found to inhibit the proliferative responses of both p195-212- and p259-271-specific T cell lines. Furthermore, the analog inhibited the in vivo priming of lymph node cells of both SJL and BALB/c mice when administered i.v., i.p., or per os. The dual analog Lys-262-Ala-207 could also immunomodulate myasthenogenic manifestations in mice with experimental autoimmune MG induced by inoculation of a pathogenic T cell line. Thus, a single peptide that is composed of analogs to two epitope specificities can be used to regulate T cell responses and disease associated with each epitope.

Amino Acid Sequence↗
PubMed1997

Protection against developmental retardation in apolipoprotein E-deficient mice by a fatty neuropeptide: implications for early treatment of Alzheimer's disease.

Stearyl-Nle17-VIP (SNV) is a novel agonist of vasoactive intestinal peptide (VIP) exhibiting a 100-fold greater potency than the parent molecule and specificity for a receptor associated with neuronal survival. Here, mice deficient in apolipoprotein E (ApoE), a molecule associated with the etiology of Alzheimer's disease, served as a model to investigate the developmental and protective effects of SNV. In comparison to control animals, the deficient mice exhibited (a) reduced amounts of VIP messenger RNA; (b) decreased cholinergic activity (c) significant retardation in the acquisition of developmental milestones: forelimb placing behavior and cliff avoidance behavior; and (d) learning and memory impairments. Daily injections of SNV to ApoE-deficient newborn pups resulted in increased cholinergic activity and marked improvements in the time of acquisition of behavioral milestones, with peptide-treated animals developing as fast as control animals and exhibiting improved cognitive functions after cessation of peptide treatment. Specificity was demonstrated in that treatment with a related peptide (PACAP), pituitary adenylate cyclase-activating peptide, produced only limited amelioration. As certain genotypes of ApoE increase the probability of Alzheimer's disease, early counseling and preventive treatments may now offer an important route for therapeutics design.

Alzheimer Disease↗
PubMed1997

Characterizing immunodominant and protective influenza hemagglutinin epitopes by functional activity and relative binding to major histocompatibility complex class II sites.

In the present study the analysis of functional activity and major histocompatibility complex (MHC) binding of two adjacent MHC class II-restricted epitopes, located in the C-terminal 306-329 region of human influenza A virus hemagglutinin 1 subunit (HA1) conserved with subtype sequences and not affected by antigenic drift, was undertaken to explore the hierarchy of local immunodominance. The functional activity of two T cell hybridomas of the memory/effector Th1 phenotype in combination with in vivo immunization studies provided a good tool for investigating the functional characteristics of the T cell response. The in vitro binding assays performed with a series of overlapping, N-terminal biotinylated peptides covering the 306-341 sequence enabled us to compare the relative binding efficiency of peptides, comprising two distinct epitopes of this region, to I-Ed expressed on living antigen-presenting cells. Our studies revealed that (i) immunization of BALB/c mice with the 306-329 H1 or H2 peptides resulted in the activation and proliferation of T cells recognizing both the 306-318 and the 317-329 epitopes, while the 306-329 H3 peptide elicits predominantly 306-318-specific T cells, (ii) the 317-329 HA1 epitope of the H1 and H2 but not the H3 sequence is recognized by T cells and is available for recognition not only in the 317-329 peptide but also in the extended 306-329 or 306-341 peptides, (iii) the 306-318 and the 317-329 hemagglutinin peptides encompassing the H1, H2 but not the H3 sequence bind with an apparently similar affinity to and therefore compete for I-Ed binding sites, and (iv) the 317-341, the 317-329 peptides and their truncated analogs show subtype-dependent differences in MHC binding and those with lower binding capacity represent the H3 subtype sequences. These results demonstrate that differences in the binding capacity of peptides comprising two non-overlapping epitopes located in the C-terminal 306-329 region of HA1 of all three subtype-specific sequences to MHC class II provide a rationale for the local and also for the previously observed in vivo immunodominance of the 306-318 region over the 317-329 epitope in the H3 but not in the H1 or H2 sequences. In good correlation with the results of the binding and functional inhibition assays, these data demonstrate that in the H1 and H2 subtypes both regions are available for T cell recognition, they compete for the same restriction element with an apparently similar binding efficiency and, therefore, function as co-dominant epitopes. Due to the stabilizing effect of the fusion peptide, peptides comprising the 306-341 or 317-341 H1 sequences are highly immunogenic and elicit a protective immune response which involves the production of antibodies and interleukin-2 and tumor necrosis factor producing effector Th1 cells both directed against the 317-329 region. Based on the similarity of the I-Ed and HLA-DR1 peptide binding grooves and motifs, these results suggest that amino acid substitutions inserted to the H3 subtype sequence during viral evolution can modify the relative MHC binding capacity and invert the local hierarchy of immunodominance of two closely situated epitopes that are able to bind to the same MHC class II molecule.

Amino Acid Sequence↗
PubMed1997

PACAP hybrid: a new PACAP receptor antagonist.

The effects of pituitary adenylate cyclase activating polypeptide (PACAP) hybrid, a synthetic antagonist, was investigated on NIH/3T3 cells containing PACAP receptor (R) splice variants (SVs). PACAPhybrid inhibited 125I-PACAP-27 binding to NIH/3T3 cells stably expressing PACAP-R basic, SV-1, SV-2 or SV-3 with an IC50 of 1000 nM. PACAPhybrid antagonized the ability of PACAP-27 to elevate cAMP regardless of the PACAP-R SV used. PACAP was more efficacious at increasing cytosolic Ca2+ in NIH/3T3 cells containing PACAP-R SV-2 than PACAP-R basic, SV-1 or SV-3. PACAPhybrid antagonized the increase in cytosolic Ca2+ caused by PACAP-27 regardless of the PACAP-R SV used. PACAP was more potent at elevating c-fos mRNA using NIH/3T3 cells transfected with PACAP-R SV-2 than PACAP-R basic, SV-1 or SV-3. PACAPhybrid antagonized the increase in c-fos mRNA caused by PACAP-27. These data suggest that PACAPhybrid is a useful PACAP receptor antagonist for PACAP-R SVs.

3T3 Cells↗
PubMed1997

(Stearyl, Norleucine17)VIP hybrid antagonizes VIP receptors on non-small cell lung cancer cells.

The effects of VIP receptor antagonists were investigated using non-small cell lung cancer (NSCLC) cells. By Northern blot and RT-PCR, VIP1 receptors were detected on NSCLC cell line NCI-H1299. VIPhybrid,(N-Stearyl-Norleucine17) VIPhybrid ((SN) VIPhybrid) and PTC4495 inhibited 125I-VIP binding to NCI-H1299 cells with IC50 values of 500, 30 and 5000 nM respectively. (SN) VIPhybrid (1 microM) had no effect on basal cAMP but strongly inhibited the increase in cAMP caused by 10 nM VIP. The order of peptide potency to inhibit cAMP was (SN) VIPhybrid > VIPhybrid > PTC4495. (SN) VIPhybrid was more potent than VIPhybrid at inhibiting NCI-H1299 colony formation. Also, (SN) VIPhybrid was more potent than VIPhybrid at inhibiting NCI-H1299 xenograft formation in nude mice. These data suggest that (SN) VIPhybrid antagonizes VIP1 receptors on NSCLC cells.

Amino Acid Sequence↗