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Biomedical subjects

J E Hunt

Publications and source records attributed to J E Hunt.

At least 19 recordsLinked to original sources
PubMed2005

The effects of UVB radiation on temperate southern hemisphere forests.

The temperate forests of the southern hemisphere are the most likely forests to be affected by increased levels of ultraviolet-B (UVB) radiation resulting from reduced ozone. The review describes these forests and then discusses the morphological changes, physiological effects, and protection mechanisms, particularly UV absorbing compounds that result from present day and increasing UVB radiation. Possible avenues for future research are explored.

Africa, Southern↗
PubMed2005

Components of ecosystem evaporation in a temperate coniferous rainforest, with canopy transpiration scaled using sapwood density.

Here we develop and test a method to scale sap velocity measurements from individual trees to canopy transpiration (E(c)) in a low-productivity, old-growth rainforest dominated by the conifer Dacrydium cupressinum. Further, E(c) as a component of the ecosystem water balance is quantified in relation to forest floor evaporation rates and measurements of ecosystem evaporation using eddy covariance (E(eco)) in conditions when the canopy was dry and partly wet. Thermal dissipation probes were used to measure sap velocity of individual trees, and scaled to transpiration at the canopy level by dividing trees into classes based on sapwood density and canopy position (sheltered or exposed). When compared with ecosystem eddy covariance measurements, E(c) accounted for 51% of E(eco) on dry days, and 22% of E(eco) on wet days. Low transpiration rates, and significant contributions to E(eco) from wet canopy evaporation and understorey transpiration (35%) and forest floor evaporation (25%), were attributable to the unique characteristics of the forest: in particular, high rainfall, low leaf area index, low stomatal conductance and low productivity associated with severe nutrient limitation.

Ecosystem↗
PubMed1999

Human tryptases alpha and beta/II are functionally distinct due, in part, to a single amino acid difference in one of the surface loops that forms the substrate-binding cleft.

Tryptases alpha and beta/II were expressed in insect cells to try to ascertain why human mast cells express these two nearly identical granule proteases. In contrast to that proposed by others, residue -3 in the propeptide did not appear to be essential for the three-dimensional folding, post-translational modification, and/or activation of this family of serine proteases. Both recombinant tryptases were functional and bound the active-site inhibitor diisopropyl fluorophosphate. However, they differed in their ability to cleave varied trypsin-susceptible chromogenic substrates. Structural modeling analyses revealed that tryptase alpha differs from tryptase beta/II in that it possesses an Asp, rather than a Gly, in one of the loops that form its substrate-binding cleft. A site-directed mutagenesis approach was therefore carried out to determine the importance of this residue. Because the D215G derivative of tryptase alpha exhibited potent enzymatic activity against fibrinogen and other tryptase beta/II-susceptible substrates, Asp215 dominantly restricts the substrate specificity of tryptase alpha. These data indicate for the first time that tryptases alpha and beta/II are functionally different human proteases. Moreover, the variation of just a single amino acid in the substrate-binding cleft of a tryptase can have profound consequences in the regulation of its enzymatic activity and/or substrate preference.

Amino Acid Sequence↗
PubMed1997

Mouse mast cell protease 9, a novel member of the chromosome 14 family of serine proteases that is selectively expressed in uterine mast cells.

Mouse mast cell protease (mMCP) 1, mMCP-2, mMCP-4, and mMCP-5 are members of a family of related serine proteases whose genes reside within an approximately 850 kilobase (kb) complex on chromosome 14 that does not readily undergo crossover events. While mapping the mMCP-1 gene, we isolated a novel gene that encodes a homologous serine protease designated mMCP-9. The mMCP-9 and mMCP-1 genes are only approximately 7 kb apart on the chromosome and are oriented back to back. The proximity of the mMCP-1 and mMCP-9 genes now suggests that the low recombination frequency of the complex is due to the closeness of some of its genes. The mMCP-9 transcript and protein were observed in the jejunal submucosa of Trichinella spiralis-infected BALB/c mice. However, in normal BALB/c mice, mMCP-9 transcript and protein were found only in those mast cells that reside in the uterus. Thus, the expression of mMCP-9 differs from that of all other chymases. The observation that BALB/c mouse bone marrow-derived mast cells developed with interleukin (IL) 10 and c-kit ligand contain mMCP-9 transcript, whereas those developed with IL-3 do not, indicates that the expression of this particular chymase is regulated by the cytokine microenvironment. Comparative protein structure modeling revealed that mMCP-9 is the only known granule protease with three positively charged regions on its surface. This property may allow mMCP-9 to form multimeric complexes with serglycin proteoglycans and other negatively charged proteins inside the granule. Although mMCP-9 exhibits a >50% overall amino acid sequence identity with its homologous chymases, it has a unique substrate-binding cleft. This finding suggests that each member of the chromosome 14 family of serine proteases evolved to degrade a distinct group of proteins.

Amino Acid Sequence↗
PubMed1996

A heparin-sensitive phospholipase A2 and prostaglandin endoperoxide synthase-2 are functionally linked in the delayed phase of prostaglandin D2 generation in mouse bone marrow-derived mast cells.

BALB/cJ mouse bone marrow-derived mast cells (BMMC) developed with interleukin (IL)-3 can be stimulated by c-kit ligand (KL) in the presence of IL-10 and IL-1beta for sequential immediate and delayed generation of prostaglandin (PG) D2 through utilization of constitutive prostaglandin endoperoxide synthase (PGHS) -1 and induced PGHS-2, respectively (Murakami, M., Matsumoto, R., Austen, K. F., and Arm, J. P. (1994) J. Biol. Chem. 269, 22269-22275). We now report that BALB/cJ BMMC stimulated with KL + IL-10 + IL-1beta also exhibit the biphasic release of [3H]arachidonic acid with an immediate phase over the first 10 min followed by a delayed phase from 2 to 7 h. The delayed phase of arachidonic acid release and of PGD2 generation was inhibited by heparin, which concomitantly released a phospholipase (PL) A2 from the cells into the supernatant. Both dexamethasone and a type II PLA2 inhibitor, 12-epi-scalaradial, suppressed delayed-phase PGD2 generation at concentrations that did not affect immediate eicosanoid generation. Transcripts for type IIA PLA2, as assessed by reverse transcription-polymerase chain reaction, were progressively induced in BALB/cJ BMMC treated for 2 to 7 h with KL + IL-10 + IL-1beta; the induction of these transcripts was down-regulated by 10(-6) M dexamethasone. The expression of steady-state transcripts and protein for cytosolic PLA2 (cPLA2) did not change. PGHS-2-dependent delayed-phase PGD2 generation elicited by IgE-dependent activation of BALB/cJ BMMC primed with KL + IL-10 was also accompanied by the induction of type IIA PLA2 transcripts and was suppressed by heparin, with concomitant release of PLA2 into the supernatant. However, both the direct, cytokine-stimulated and the cytokine-primed, IgE-dependent, delayed-phase PGD2 generation occurred in BMMC from C57BL/6J mice, which have a natural disruption of the type IIA PLA2 gene. Thus, kinetic, pharmacologic, and genetic analyses suggest that an inducible, heparin-sensitive PLA2, rather than cPLA2, provides arachidonic acid to concomitantly induced PGHS-2 for delayed-phase PGD2 biosynthesis in activated BMMC. Furthermore, this heparin-sensitive PLA2 likely represents a novel PLA2 or a new function for a known low molecular weight PLA2.

Animals↗
PubMed1996

Natural disruption of the mouse mast cell protease 7 gene in the C57BL/6 mouse.

The C57BL/6 mouse differs from the BALB/c mouse in that its ear and skin mast cells and its progenitor bone marrow-derived mast cells (mBMMCs) do not express mouse mast cell protease (mMCP) 7. We now report that, as detected by nuclear run-on analysis, the mMCP-7 gene is transcribed in C57BL/6 mBMMCs at a rate comparable to that in BALB/c mBMMCs. Reverse transcriptase-polymerase chain reaction analysis and sequencing of the product revealed that the ears of C57BL/6 mice contain small amounts of a mMCP-7 transcript that possesses a 98-base pair deletion. The deletion begins at a normally quiescent cryptic splice site (G416TGAG), 98 base pairs upstream of the normal exon 2/intron 2 splice site (G514TGAG), and introduces a premature stop codon in the alternatively spliced transcript. Thus, even if translated, the mature protein would consist of only 18 amino acids as compared to 245 amino acids in normal mMCP-7. Sequence analysis of the mMCP-7 gene in the C57BL/6 mouse revealed that the cryptic splice site is activated due to a G514-->A point mutation at the first nucleotide of the normal exon 2/intron 2 splice site. This is the first report of a mutation of a gene that encodes a mast cell secretory granule constituent that leads to its loss of expression. Moreover, the mMCP-7 gene is the first found in any species that sequentially has undergone a splice site mutation to cause retention of an intron and then a second splice site mutation to cause activation of a cryptic splice site.

Amino Acid Sequence↗
PubMed1995

Recognising promoter sequences using an artificial immune system.

We have developed an artificial immune system (AIS) which is based on the human immune system. The AIS possesses an adaptive learning mechanism which enables antibodies to emerge which can be used for classification tasks. In this paper, we describe how the AIS has been used to evolve antibodies which can classify promoter containing and promoter negative DNA sequences. The DNA sequences used for teaching were 57 nucleotides in length and contained procaryotic promoters. The system classified previously unseen DNA sequences with an accuracy of approximately 90%.

Algorithms↗
PubMed1994

Fullerenes in the 1.85-billion-year-old Sudbury impact structure.

Fullerenes (C60 and C70) have been identified by laser desorption, laser desorption post-ionization, and high-resolution electron-impact mass spectrometry in shock-produced breccias (Onaping Formation) of the Sudbury impact structure in Ontario, Canada. The C60 isotope is present at a level of a few parts per million. The fullerenes were likely synthesized within the impact plume from the carbon contained in the bolide. The oxidation of the fullerenes during the 1.85 billion years of exposure was apparently prevented by the presence of sulfur in the form of sulfide-silicate complexes associated with the fullerenes.

Carbon↗
PubMed1994

Sneddon's syndrome, anti-cardiolipin antibody and glomerular thrombosis.

Sneddon's syndrome, cerebrovascular thrombosis and livedo reticularis, is often a variant of the "primary" anti-phospholipid syndrome (PAPS). We report a woman with PAPS, presenting as Sneddon's syndrome, with renal impairment and glomerular thrombosis on renal biopsy. An IgG anti-cardiolipin antibody (aCL) was identified. The aCL was purified by affinity chromatography, gel filtration chromatography and ion-exchange chromatography, assayed in a modified ELISA and found to be of the type that requires the plasma protein beta 2-GPI to bind aCL. As beta 2-GPI has anticoagulant properties it is postulated that its interaction with aCL has a pathogenic role in the thrombotic lesions associated with aCL.

Adult↗
PubMed1993

Identification of a region of beta 2-glycoprotein I critical for lipid binding and anti-cardiolipin antibody cofactor activity.

beta 2-Glycoprotein I (beta 2-GPI), a phospholipid-binding plasma protein, is an absolute requirement (cofactor) for the binding of autoimmune-type anti-cardiolipin (aCL) antibodies to cardiolipin (CL). The nature of this cofactor activity and the specific regions of the molecule involved have not yet been determined. We have identified a preparation of beta 2-GPI that lacks aCL antibody cofactor activity. Analysis of the structural differences between the active and inactive forms enabled identification of the region of beta 2-GPI critically important for aCL cofactor activity. The active form of beta 2-GPI bound CL and displayed cofactor activity down to 1 microgram/ml. The inactive form failed to bind CL and possessed no cofactor activity even at concentrations up to 94 micrograms/ml, indicating that the ability of beta 2-GPI to bind lipids is an absolute requirement for aCL cofactor activity. Both forms possessed identical N-terminal sequences and were recognized as essentially immunoreactively identical by polyclonal antisera to beta 2-GPI. However, the inactive form has undergone proteolytic cleavage and exists primarily as a "clipped" molecule, the polypeptide chain being cleaved between Lys-317 and Thr-318 (a potential thrombin cleavage site), with the two cleaved segments linked as a disulfide-bonded complex. This indicates that the C-terminal region is critically important for beta 2-GPI to bind lipid and for aCL cofactor activity. The clipped form of beta 2-GPI would not be suitable for use as aCL cofactor and its use may have led some investigators to conclude incorrectly that beta 2-GPI does not interact with aCL antibodies.

Amino Acid Sequence↗
PubMed1993

Anterior translocation of the tibia at MR imaging: a secondary sign of anterior cruciate ligament tear.

The authors evaluated measurement of the degree of anterior subluxation ("translocation") of the tibia in regard to the femur as a predictor of anterior cruciate ligament (ACL) tear. Eighty-nine magnetic resonance (MR) imaging studies of patients with either an arthroscopically confirmed intact (n = 29), acutely torn (n = 27), or chronically torn (n = 33) ACL were retrospectively reviewed. The degree of translocation was measured on hard-copy images by using two methods. Buckling of the posterior cruciate ligament (PCL) was also evaluated. Anterior tibial translocation, when measured at the midsagittal plane of the lateral femoral condyle with regard to a plane parallel to the cephalocaudal axis of the image, was a relatively specific indicator of ACL disruption. Subluxation of 5 mm or more had 58% sensitivity, 93% specificity, and 69% accuracy for an ACL tear. All knees with subluxation of 7 mm or more had torn ACLs. Buckling of the PCL was less sensitive and less accurate than anterior translocation as an indicator of ACL disruption.

Anterior Cruciate Ligament↗
PubMed1993

Immunohistochemical reactivity of basal and luminal epithelium-specific cytokeratin antibodies within normal and neoplastic canine mammary glands.

Human basal epithelium (myoepithelium)-specific (312C8-1) and luminal epithelium-specific (13H5) cytokeratin antibodies were applied to frozen sections of normal canine mammary tissues (seven), benign adenomas and hyperplasias (five), mixed tumors (12), and adenocarcinomas (18) to determine if epithelial subsets could be discriminated by the use of an avidin biotin peroxidase complex immunohistochemical procedure. The 312C8-1 and 13H5 antibodies were consistently reactive with basal and luminal epithelium, respectively, in the normal mammary gland (7/7) and in benign adenomas and hyperplasias (5/5). Mixed mammary tumors had similar basal and luminal epithelial reactivity and also had proliferating spindle-shaped stromal cells that were reactive with 312C8-1 (10/12) and 13H5 (4/12). The adenocarcinomas were subclassified into basal, luminal, and basal/luminal on the basis of 312C8-1 reactivity (4/18), 13H5 reactivity (2/18), and dual reactivity with mutually exclusive anatomic distribution (11/18), respectively. Those tumors with dual immunoreactivity were indicative of noninvasive carcinomas. Dogs with neoplasms that were reactive with 312C8-1 and nonreactive with 13H5 had local recurrence or distant metastasis within 2 weeks to 6 months after diagnosis. Other antibodies used for comparison were pan cytokeratin AE1/AE3, actin HHF35, and vimentin. 312C8-1 and 13H5 antibodies are specific for canine mammary basal and luminal epithelium, respectively, and by employing these antibodies, the origin and differentiation of canine mammary neoplasms can be determined more accurately than on the basis of hematoxylin and eosin-stained tissue alone.

Actins↗
PubMed1993

Antibodies to phosphatidylethanolamine in antiphospholipid syndrome and systemic lupus erythematosus: their correlation with anticardiolipin antibodies and beta 2 glycoprotein-I plasma levels.

The sera patients with systemic lupus erythematosus (SLE) and antiphospholipid syndrome (APS) were tested, by ELISA, for antibodies to phosphatidylethanolamine (aPE), as well as to cardiolipin (aCL) and compared to healthy blood donors (HBD). Both, SLE and APS patients presented a higher titre of IgM-aPE antibodies than normals, while the IgG and IgA aPE reactivity did not differ. APS patients were characterized by higher IgM-aPE antibody titres than SLE patients. In contrast, the predominant isotype of aCL antibodies in APS patients was IgG. The IgM aPE reactivity was correlated with IgM aCL reactivity, while no correlation was observed between the total IgM values and IgM-aPE binding units of sera tested. Since it was shown that beta 2-glycoprotein-I (beta 2-GPI) contributes to a complex antigen by binding to phospholipids and that this antigen is recognized by antiphospholipid antibodies from autoimmune patients, sera beta 2-GPI levels were measured and correlated to aCL and APE activity. Although APS patients had higher beta 2-GPI levels than SLE patients, no correlation was found between the beta 2-GPI levels and IgG/IgM aCL and IgM-aPE reactivities a finding suggesting that in addition to beta 2-GPI, other cofactors for aPE antibodies may exist. These findings indicate that aPE and aCL antibodies co-exist and that the IgM-isotype is predominant in APS. In addition, the IgA and IgG aPE antibodies appear to occur in low titres in these patients, as well as in normals and may exist as natural autoantibodies. We suggest that the high IgM-aPE antibodies may be viewed as a thymus independent process.

Antibodies, Anticardiolipin↗
PubMed1992

Patients with anticardiolipin antibodies with and without antiphospholipid syndrome: their clinical features and beta 2-glycoprotein-I plasma levels.

The clinical and serological features of 38 aCL-positive patients were compared to those of 45 aCL-negative patients. A significantly higher incidence of thrombophlebitis and livedo reticularis was found in aCL-positive patients. There were 13 aCL positive patients with thrombophlebitis and/or arterial thromboses and these 13 patients were designated as having the antiphospholipid syndrome (APS) while the remaining 70 patients were diagnosed as having Systemic Lupus Erythematosus (SLE). APS patients also had a high incidence of arterial occlusions, recurrent abortions and strokes compared to SLE patients. Patients with high levels of IgG-aCL were more likely to have APS, while patients with low levels of IgG-aCL or IgM-aCL only were more likely to have SLE without the clinical features of APS. Since aCL antibodies have recently been shown to interact with a phospholipid-binding plasma protein beta 2-glycoprotein-I (beta 2-GPI), we measured the beta 2-GPI levels in these patients and found that beta 2-GPI levels are significantly higher in APS compared to SLE patients negative for aCL antibodies. Since beta 2-GPI is known to exert multiple effects on coagulation processes the interaction of aCL antibodies with this glycoprotein may play a pathogenic role in APS.

Antiphospholipid Syndrome↗
PubMed1992

A phospholipid-beta 2-glycoprotein I complex is an antigen for anticardiolipin antibodies occurring in autoimmune disease but not with infection.

Anticardiolipin antibodies (aCL) purified from patients with autoimmune disease have recently been shown to interact with a phospholipid-binding plasma protein, beta 2-glycoprotein I (beta 2-GPI). The aim of this study was to determine whether aCL purified from patients with infection also interact with beta 2-GPI. aCL purified from 23 patients with malaria, infectious mononucleosis, tuberculosis, hepatitis A or syphilis did not require the presence of beta 2-GPI to bind cardiolipin (CL). In contrast, aCL were purified from 11 out of 12 patients with autoimmune disease that bound CL only in the presence of beta 2-GPI. Thrombotic complications appear to be associated with aCL occurring in autoimmune disease but not with aCL associated with infections. We postulate that this increased risk of thrombosis in the autoimmune group may be due to the presence of aCL that bind CL in association with beta 2-GPI, a plasma protein with anticoagulant activity.

Antibodies, Anticardiolipin↗