Polygalacturonase inhibiting protein in plant cell walls

Polygalacturonase inhibiting protein (PGIP) is localized in plant cell walls and plays an important role both in pectic substance metabolism and in prevention of the penetration of phytopathogenic microorganisms. Apparently, PGIP is responsible for the specificity of cell--cell interactions during pollination or inoculation by fungi nonpathogenic for the particular plant. PGIPs from different plants share a basic common structure. They are rather thermostable glycoproteins enriched with leucine and contain about 20% carbohydrates; the molecular weight varies between 37-54 kD. The synthesis of PGIP is encoded by one gene, and its expression is stimulated by injury and fungal infection. The resistance of plant tissues to infection frequently correlates with PGIP expression and with inhibiting action on fungal PG. Thus, PGIP is believed to be useful for gene engineering to obtain transgenic plants resistant to fungal infection or retaining commercial value during storage.     

Microbial succession and intestinal enzyme activities in the developing rat

J. CHANG, R.W. CHADWICK, J.C. ALLISON, Y.O. HAYES, D.L. TALLEY AND C.E. AUTRY. 1994. The succession of gut bacteria and selected intestinal enzyme activities in developing 7–35-d-old rats was studied. Aerobes and anaerobes were identified as members of four broad major bacterial groups, i.e. Gram-positive rods, Gram-positive cocci, Gram-negative rods and obligate anaerobes. The enzyme activities of nitro and azo reductases, β-glucuronidase, dechlorinase and dehydrochlorinase were determined by anaerobic incubation of intestinal homogenates with 3,4-dichloronitrobenzene, methyl orange, ***p-nitrophenyl-β-D-glucuronide, and ***p, p-DDT respectively. Nitroreductase and azo reductase activities increased significantly with the appearance of anaerobes in the large intestine. No increase in either nitroreductase or azo reductase activities in the small intestine was found. The early and high level of β-glucuronidase activity in the small and large intestines coincided with high numbers of coliforms recovered in 7 and 14 d animals. Dehydrochlorinase activity appeared early but was undetectable at both 21 and 28 d. Its activity increased at 35 d. Dechlorinase activity was variable in development. The rapid changes in the microbial flora and intestinal enzyme activities may influence the susceptibility of pre-pubescent rats to a variety of toxicants. Therefore, age-dependent toxicity may be important in the risk assessment of some environmental chemicals.     

Ichthyofaunal utilization of newly-created versus natural salt marsh creeks in Mission Bay,CA

Loss of wetland habitat has proceeded at an alarmingrate in southern California, and increasingly marshrestoration and creation are being used to mitigatethese losses. As part of an effort to evaluatefunctional equivalence of created systems, theichthyofaunal assemblages in a created and adjacentnatural marsh in Mission Bay, San Diego, Californiawere compared. Fishes trapped in both marshes includedFundulus parvipinnis, Gillichthysmirabilis, Acanthogobius flavimanus, Ctenogobius sagittula, Atherinops affinis, andMugil cephalus. Fundulus parvipinniswasnumerically dominant in both systems, representing onaverage 69% of all fishes trapped in the createdmarsh and 65% of all fishes trapped in the naturalmarsh. Gillichthys mirabiliswas the second-mostabundant species, representing on average 31% of allfishes trapped in the created marsh and 28% of allfishes trapped in the natural marsh. Species richnessand dominance measures were similar between the twosystems, while abundances were higher in the naturalrelative to the created marsh. The size-structure ofF. parvipinnisand G. mirabilisdifferedbetween the created and natural marsh creeks, with thecreated marsh populations being skewed towards largersize classes. These size differences are believed toarise from differences in creek morphology between thecreated and natural systems, and potentially affectboth predators and prey of these species in the marsh.Mark-release-recapture revealed considerable marshfidelity, with as many as 35% of the F.parvipinnistagged in a marsh being recovered one daylater in the same marsh. Stable isotope analyses ofF. parvipinnisrevealed similar ¹⁵Nand ³⁴S values between marshes; howeverthere was a consistent enrichment in ¹³C (>3per mil) in tissues of F. parvipinnisfrom thecreated marsh, supporting the high marsh fidelitysuggested by tagging results. This first publisheddocumentation of the Mission Bay marsh resident fishessuggests that the created marsh ichthyofaunalassemblage was distinct in density and size structurefrom the adjacent natural marsh, and provides lessonsfor future restoration efforts.     

Relationship of gastric emptying and volume changes after a solid meal in humans

Noninvasive imaging has been developed to measure gastric volumes. The relationship between gastric emptying and volume postprandially is unclear. The aims were to 1) develop a 3-dimensional (3D) single photon emission-computed tomography (SPECT) method to simultaneously measure gastric volume and emptying postprandially, 2) describe the course of gastric volume change during emptying of the meal, and 3) assess a 3D method measuring gastric emptying. In 30 healthy volunteers, we used (111)In-planar and (99m)Tc-SPECT imaging to estimate gastric emptying and volume after a radiolabeled meal. A customized analysis program of SPECT imaging assessed gastric emptying. A Bland-Altman plot assessed the performance of the new SPECT analysis compared with planar analysis. Gastric volume postprandially exceeds the fasting volume plus meal volume. The course of volume change and gastric emptying differ over time. Higher differences in volumes exist relative to fasting plus residual meal volumes at 15 min (median 763 vs. 568 ml, respectively, P < 0.001), 1 h (median 632 vs. 524 ml, P < 0.001), and 2 h (median 518 vs. 428 ml, P < 0.02), in contrast to similar volumes at 3 h (median 320 vs. 314 ml, P = 0.85). Analysis of SPECT imaging accurately measures gastric emptying compared with planar imaging with median differences of 1% (IQR -2.25 to 2.0) at 1 h, 1% (-3.25 to 2.25) at 2 h, and -2.5% (-4 to 0) at 3 h. Gastric volume exceeds meal volume during the first 2 postprandial hours, and simultaneous measurements of gastric volume and emptying can be achieved with a novel 3D SPECT method.     

Antibodies to citrullinated proteins and differences in clinical progression of rheumatoid arthritis

Antibodies to citrullinated proteins (anti-cyclic-citrullinated peptide [anti-CCP] antibodies) are highly specific for rheumatoid arthritis (RA) and precede the onset of disease symptoms, indicating a pathogenetic role for these antibodies in RA. We recently showed that distinct genetic risk factors are associated with either anti-CCP-positive disease or anti-CCP-negative disease. These data are important as they indicate that distinct pathogenic mechanisms are underlying anti-CCP-positive disease or anti-CCP-negative disease. Likewise, these observations raise the question of whether anti-CCP-positive RA and anti-CCP-negative RA are clinically different disease entities. We therefore investigated whether RA patients with anti-CCP antibodies have a different clinical presentation and disease course compared with patients without these autoantibodies. In a cohort of 454 incident patients with RA, 228 patients were anti-CCP-positive and 226 patients were anti-CCP-negative. The early symptoms, tender and swollen joint count, and C-reactive protein level at inclusion, as well as the swollen joint count and radiological destruction during 4 years of follow-up, were compared for the two groups. There were no differences in morning stiffness, type, location and distribution of early symptoms, patients' rated disease activity and C-reactive protein at inclusion between RA patients with and without anti-CCP antibodies. The mean tender and swollen joint count for the different joints at inclusion was similar. At follow-up, patients with anti-CCP antibodies had more swollen joints and more severe radiological destruction. Nevertheless, the distribution of affected joints, for swelling, bone erosions and joint space narrowing, was similar. In conclusion, the phenotype of RA patients with or without anti-CCP antibodies is similar with respect to clinical presentation but differs with respect to disease course.     

Fluorescence quenching analysis of the association and dissociation of a diarylheterocycle to cyclooxygenase-1 and cyclooxygenase-2: dynamic basis of cyclooxygenase-2 selectivity

Cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) are the enzymes responsible for the biosynthesis of the precursor to the biologically active prostaglandins, prostacyclin, and thromboxane and are the molecular targets for nonsteroidal antiinflammatory drugs (NSAIDs). Selective COX-2 inhibitors are antiinflammatory and analgesic but lack gastrointestinal toxicity, an undesirable side effect attributed to COX-1 inhibition. Crystallographic analysis of selective COX inhibitors complexed with either isoform provides some information about the molecular determinants of selectivity but does not provide information about the dynamics of inhibitor association/dissociation. We employed rapid-mixing techniques and fluorescence quenching to monitor the association and dissociation of a selective COX-2 inhibitor to COX-1 or COX-2. The association of the fluorescent diaryloxazole, SC299, with both enzymes occurs in a time-dependent fashion. Its binding to COX-2 occurs in three kinetically distinct steps whereas its binding to COX-1 occurs in two steps. In contrast to the relatively rapid association of SC299 with both enzymes, its dissociation from COX-2 is quite slow and occurs over several hours whereas the dissociation from COX-1 is complete in less than 1 min. The selectivity of SC299 as a COX-2 inhibitor correlates to its relative rates of dissociation from the two COX isoforms. A model is proposed for diarylheterocycle binding to COX's that integrates these kinetic data with available structural information.     

A Quantitative MRI Method for Imaging Blood-Brain Barrier Leakage in Experimental Traumatic Brain Injury

Blood-brain barrier (BBB) disruption is common following traumatic brain injury (TBI). Dynamic contrast enhanced (DCE) MRI can longitudinally measure the transport coefficient K^trans which reflects BBB permeability. K^trans measurements however are not widely used in TBI research because it is generally considered to be noisy and possesses low spatial resolution. We improved spatiotemporal resolution and signal sensitivity of K^trans MRI in rats by using a high-sensitivity surface transceiver coil. To overcome the signal drop off profile of the surface coil, a pre-scan module was used to map the flip angle (B₁ field) and magnetization (M₀) distributions. A series of T₁-weighted gradient echo images were acquired and fitted to the extended Kety model with reversible or irreversible leakage, and the best model was selected using F-statistics. We applied this method to study the rat brain one hour following controlled cortical impact (mild to moderate TBI), and observed clear depiction of the BBB damage around the impact regions, which matched that outlined by Evans Blue extravasation. Unlike the relatively uniform T₂ contrast showing cerebral edema, K^trans shows a pronounced heterogeneous spatial profile in and around the impact regions, displaying a nonlinear relationship with T₂. This improved K^trans MRI method is also compatible with the use of high-sensitivity surface coil and the high-contrast two-coil arterial spin-labeling method for cerebral blood flow measurement, enabling more comprehensive investigation of the pathophysiology in TBI.