Biomedical subjects

P Amiri

Publications and source records attributed to P Amiri.

11 recordsLinked to original sources

PubMed1998

Early activation of c-Jun N-terminal kinase and p38 kinase regulate cell survival in response to tumor necrosis factor alpha.

Fas ligand and tumor necrosis factor alpha (TNF) bind to members of the TNF receptor superfamily. Stimulation by Fas ligand results in apoptosis, whereas TNF induces multiple effects including proliferation, differentiation, and apoptosis. Activation of the c-Jun N-terminal kinase (JNK) and p38 kinase pathways is common to Fas and TNF signaling; however, their role in apoptosis is controversial. Fas receptor cross-linking induces apoptosis in the absence of actinomycin D and activates JNK in a caspase-dependent manner. In contrast, TNF requires actinomycin D for apoptosis and activates JNK and p38 kinase with biphasic kinetics. The first phase is transient, precedes apoptosis, and is caspase-independent, whereas the second phase is coincident with apoptosis and is caspase-dependent. Inhibition of early TNF-induced JNK and p38 kinases using MKK4/MKK6 mutants or the p38 inhibitor SB203580 increases TNF-induced apoptosis, whereas expression of wild type MKK4/MKK6 enhances survival. In contrast, the Mek inhibitor PD098059 has no effect on survival. These results demonstrate that early activation of p38 kinase (but not Mek) are necessary to protect cells from TNF-mediated cytotoxicity. Thus, early stress kinase activation initiated by TNF plays a key role in regulating apoptosis.

Amino Acid Chloromethyl Ketones↗

PubMed1996

Requirement of an ICE-like protease for induction of apoptosis and ceramide generation by REAPER.

Genetic studies indicated that the Drosophila melanogaster protein REAPER (RPR) controls apoptosis during embryo development. Induction of RPR expression in Drosophila Schneider cells rapidly stimulated apoptosis. RPR-mediated apoptosis was blocked by N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (Z-VAD-fmk), which suggests that an interleukin-1 beta converting enzyme (ICE)-like protease is required for RPR function. RPR-induced apoptosis was associated with increased ceramide production that was also blocked by Z-VAD-fmk, which suggests that ceramide generation requires an ICE-like protease as well. Thus, the intracellular RPR protein uses cell death signaling pathways similar to those used by the vertebrate transmembrane receptors Fas (CD95) and tumor necrosis factor receptor type 1.

Amino Acid Chloromethyl Ketones↗

PubMed1994

Anti-immunoglobulin E treatment decreases worm burden and egg production in Schistosoma mansoni-infected normal and interferon gamma knockout mice.

The immunoglobulin E (IgE) response is generally considered an essential component of the host defense against parasitic helminths such as Schistosoma mansoni. In contrast, work on antischistosome vaccines suggests that interferon gamma (IFN-gamma) is the critical immune mediator for vaccine-induced immunity to the parasite. In this study, the total IgE response to a primary S. mansoni infection was suppressed by anti-IgE treatment in both normal mice and in mice with defective IFN genes (gene knockout [GKO]). Reduction of the IgE response resulted in decreased worm burden and a decrease in the number of eggs produced per worm in both normal and GKO mice. Whereas anti-IgE treatment also resulted in reduced hepatosplenomegaly, granulomas around existing schistosome eggs showed normal cellularity. Serum interleukin 4 levels fell in response to the reduction in serum IgE as well. The data suggest that IgE plays a detrimental, rather than beneficial, role for the host in schistosomiasis. Furthermore, the absence of IFN-gamma was found to be of little consequence to the host-response to adults or eggs in a primary schistosome infection.

Animals↗

PubMed1992

Tumour necrosis factor alpha restores granulomas and induces parasite egg-laying in schistosome-infected SCID mice.

Schistosomiasis (bilharzia) is a parasitic disease caused by several species of schistosome worms (blood flukes). The key pathogenic event in this disease is the formation of granulomas around schistosome eggs trapped in portal venules of the liver. Granulomas are a distinctive form of chronic inflammation characterized by localized aggregation of activated macrophages around an inciting stimulus. Each granuloma evolves to form a fibrous scar; in schistosomiasis, the result is widespread hepatic fibrosis and portal hypertension. To identify the specific immune signal molecules necessary for granuloma formation, we studied schistosome infections in severe combined immunodeficient (SCID) mice, which have normal macrophages but lack functional B or T lymphocytes. Here we report that the immunoregulatory cytokine tumour necrosis factor alpha is necessary and sufficient to reconstitute granuloma formation in schistosome-infected SCID mice. Moreover, we find that the parasitic worms require tumour necrosis factor alpha for egg-laying and for excretion of eggs from the host. The implication of this latter result is that the parasite has adapted so successfully to its host that it uses a host-derived immunoregulatory protein as a signal for replication and transmission.

Animals↗

PubMed1992

Betaine:homocysteine methyltransferase from rat liver: purification and inhibition by a boronic acid substrate analog.

Betaine:homocysteine methyltransferase (BHMT) from rat liver has been highly purified by an efficient procedure requiring only two chromatographic steps: Sephadex G-100 chromatography and fast protein liquid chromatography chromatofocusing. A 170-fold purification and 7.5% overall yield were achieved. Chromatofocusing yielded three active forms of BHMT with pI values near 8.0, 7.6, and 7.0. The subunit molecular weight of each active form is 45,000 Da as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and the native enzyme has a molecular weight of 270,000 as determined by exclusion chromatography. The stability of the purified enzyme was found to be potentiated by the presence of 1 mM dimethylglycine and 1 mM homocysteine. Boronate analogs of betaine (pinanyl N,N,N-trimethylaminomethaneboronate) (4) and dimethylglycine (pinanyl N,N-dimethylaminomethaneboronate) were synthesized from pinanyl iodomethaneboronate (3) and trimethylamine or dimethylamine, respectively. The free acid of the betaine analog (5) was reversibly generated from (4). The inhibition of BHMT by (5) appears competitive with a Ki = 45 microM. Since the Km for betaine measured with the purified enzyme is near 0.1 mM, the boronic acid analog of betaine appears to function effectively as a substrate analog inhibitor of BHMT. The analog does not appear to act as a methyl donor to homocysteine when (5) is substituted for betaine in the enzyme reaction. In addition, an enzyme assay based upon C3-cyano reverse phase HPLC detection of the o-phthalaldehyde derivative of methionine was developed as an alternative to the standard radiochemical assay. Betaine:homocysteine methyltransferase in the picomole range can be quantitated using this assay as indicated by a linear response of enzyme activity to protein concentration.

Animals↗

PubMed1992

Schistosoma mansoni: cell-specific expression and secretion of a serine protease during development of cercariae.

Eukaryotic serine proteases are an important family of enzymes whose functions include fertilization, tissue degradation by neutrophils, and host invasion by parasites. To avoid damaging the cells or organisms that produced them, serine proteases must be tightly regulated and sequestered. This study elucidates how the parasitic blood fluke Schistosoma mansoni synthesizes, stores, and releases a serine protease during differentiation of its invasive larvae. In situ hybridization with a cDNA probe localized the protease mRNA to acetabular cells, the first morphologically distinguishable parasite cells that differentiate from the embryonic cell masses present in the intermediate host snail. The acetabular cells contained vimentin but not cytokeratins, consistent with a mesenchymal, not epithelial, origin. Antiprotease antibodies, localized by immunoperoxidase, showed that the protease progressively accumulated in these cells and was packaged in vesicles of three morphologic types. Extension of cytoplasmic processes containing protease vesicles formed "ducts" which reached the anterior end of fully differentiated larvae. During invasion of human skin, groups of intact vesicles were released through the acetabular cytoplasmic processes and ruptured within the host tissue. Ruptured protease vesicles were noted adjacent to degraded epidermal cells and dermal-epidermal basement membrane, as well as along the surface of the penetrating larvae themselves. These observations are consistent with the proposed dual role for the enzyme in facilitating invasion of host skin by larvae and helping to release the larval surface glycocalyx during metamorphosis to the next stage of the parasite.

Animals↗

PubMed1992

Ca(2+)-regulated serine protease associated with the nuclear scaffold.

The nuclear scaffold (NS) is a proteinaceous network of orthogonally arrayed intermediate filament proteins, termed lamins, which is responsible for nuclear structure. Recent work has demonstrated that a subset of lamins A/C is proteolytically cleaved to produce an ATP-binding protein. This proteolytic cleavage is accomplished by a NS protease activity, which shows a considerable selectivity for lamins A/C and is stringently regulated by Ca2+ in vitro, suggesting that it might also participate in control of NS breakdown in various scenarios. Here, we identify the major NS protease as a novel serine protease with a predominantly chymotryptic-like substrate preference, and we show that even transient perturbations in cytosolic Ca2+ have significant effects on the NS protease activity. This NS protease activity shows extensive similarities to the multicatalytic proteinase complex. In addition to a potential role in control of NS breakdown at mitosis and/or under pathological conditions, this NS protease is also strategically located for other functions, such as inactivation of various oncogenic proteins or maturation-promoting factor.

Amino Acid Sequence↗

PubMed1991

Arresting tissue invasion of a parasite by protease inhibitors chosen with the aid of computer modeling.

Computer modeling of the three-dimensional structure of an enzyme, based upon its primary sequence alone, is a potentially powerful tool to elucidate the function of enzymes as well as design specific inhibitors. The cercarial (larval) protease from the blood fluke Schistosoma mansoni is a serine protease hypothesized to assist the schistosome parasite in invading the human circulatory system via the skin. A three-dimensional model of the protease was built, taking advantage of the similarity of the sequence of the cercarial enzyme to the trypsin-like class of serine proteases. A large hydrophobic S-1 binding pocket, suspected from previous kinetic studies, was located in the model and confirmed by new kinetic studies with both synthetic peptide substrates and inhibitors. Unexpected structural characteristics of the enzyme were also predicted by the model, including a large S-4 binding pocket, again confirmed by assays with synthetic peptides. The model was then used to design a peptide inhibitor with 4-fold increased solubility, and a series of synthetic inhibitors were tested against live cercariae invading human skin to confirm that predictions of the model were also applicable in a biologic assay.

Amino Acid Sequence↗

PubMed1988

The Schistosomatium douthitti cercarial elastase is biochemically and structurally distinct from that of Schistosoma mansoni.

The cercarial acetabular gland proteinase of Schistosomatium douthitti, an agent of 'swimmer's itch', has been identified and characterized. Like the corresponding proteinase of Schistosoma mansoni, it has significant elastase activity and can degrade a model of dermal extracellular matrix. However, unlike the S. mansoni enzyme, it has a higher molecular weight (50,000 versus 30,000), is of a different proteinase class (metallo versus serine), and has no significant primary structure homology to the S. mansoni proteinase. While these findings indicate that the failure of S. douthitti to produce chronic schistosomiasis in humans is not due to its lacking, or having a less potent 'penetration proteinase' than S. mansoni, the proteolytic enzymes are sufficiently different to support the hypothesis that the Schistosomatium line diverged quite early from the main branch of Schistosoma evolution.

Animals↗

PubMed1984

Benzamidomethaneboronic acid: synthesis and inhibition of chymotrypsin.

Benzamidomethaneboronic acid (2) has been synthesized unambiguously from the reaction of dibutyl iodomethaneboronate and N-lithiohexamethyldisilazane to form dibutyl [bis(trimethylsilyl)amino]methaneboronate (4), which was desilylated, benzoylated, and hydrolyzed to 2. It has been shown that 2 is a strong competitive inhibitor of alpha-chymotrypsin (Ki = 8.1 X 10(-6) M, pH 7.5). The reaction product from dibutyl iodomethaneboronate and sodiobenzamide, previously shown to be a potent inhibitor of chymotrypsin, was shown by this work to be O-linked isomer, benzimidoxy-methaneboronic acid (3). The pH-Ki profile over the pH range 6.5-9.5 was consistent with the formation of an enzyme-inhibitor complex which resembled the metastable tetrahedral reaction intermediates occurring during acylation and deacylation of chymotrypsin-catalyzed hydrolysis.

Acylation↗

PubMedDate not supplied

Experimental induction of caprylate-dependent albumin antibodies.

To provide evidence for the immune nature of the albumin agglutination phenomenon or caprylate dependent albumin agglutinins (CDAA), rabbits were injected with native serum that had been incubated in a solution of sodium caprylate. Two of three rabbits responded with the production of CDAA, which in vitro behaved identical to human antisera. Human cells were agglutinated only when caprylate stabilized albumin was added to the cell/serum mixture or when caprylate free albumin plus sodium caprylate were used. The CDAA failed to agglutinate rabbit cells, although, both human adult and cord cells were agglutinated. These experiments indicate that the CDAA represent an immune response to native albumin that had been altered by caprylate.

Agglutination↗