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Biomedical subjects

J R Pratt

Publications and source records attributed to J R Pratt.

At least 19 recordsLinked to original sources
PubMed1997

Developmental regulation of the plant mitochondrial matrix located HSP70 chaperone and its role in protein import.

Changes in the level of the mitochondrial chaperone mtHSP70 have been investigated in pea (Pisum sativum) leaf mitochondria by Western blot analysis and quantified by scanning densitometry. As pea leaves develop (from 6 days to 30 days of age) the levels of mtHSP70 decrease. Analysis of the levels of the alpha subunit of the F1ATPase show that the levels of this protein remain constant throughout the same developmental period, whereas the levels of the alternative oxidase increase. In vitro import of the alternative oxidase precursor protein into pea leaf mitochondria from day 6 to day 30 leaves and quantification by scanning densitometry indicates that protein import efficiency decreases with increasing maturity of the plant cell. Results are discussed in terms of how changing levels of the mtHSP70 chaperone, as a result of plant cell development, influence the efficiency of protein import.

Blotting, Western↗
PubMed1997

Influence of complement on the allospecific antibody response to a primary vascularized organ graft.

The induction of antibody responses against T cell-dependent antigens has been reported to be influenced by complement. We therefore asked if the primary induction of alloantibodies against transplantation antigens, an important determinant of transplant outcome, is complement sensitive and whether this has functional implications. We transplanted rat kidney allografts into fully major histocompatibility complex-mismatched recipients, in which complement activation was inhibited by daily injection of soluble recombinant human complement receptor type 1 (sCR1). Control allograft recipients were injected with saline. Animals in the control group showed a marked antibody response against donor-specific antigens and an increase in the proportion of activated B and T splenocytes by day 5 after transplantation. Complement-inhibited rats showed a reduced level of antibody binding on target cells sharing the same histocompatibility antigens as the donor strain (p < 0.001), and a reduced level of activated splenic B (p < 0.01) and T (p < 0.01) cells. In a functional assay, the plasma of complement-inhibited rats showed reduced cytotoxic activity against donor-specific cells, and their grafts contained less bound antibody than controls. Analysis beyond 6 days was obscured due to the development of antibodies against sCR1. We conclude that complement activation facilitates the induction of the alloantibody response. Sparing of vascular injury and prolongation of graft survival, previously reported in complement-inhibited rats (Pratt J. R. et al., Am. J. Path. 1996, 149: 2055), could therefore be due to down-regulation of the B cell response as well as reduced complement-dependent cytotoxicity. Inhibition of complement may provide an ancillary approach to the prevention of allospecific antibody formation and the prolongation of allograft survival in primary kidney grafting.

Animals↗
PubMed1997

Expression of tissue type plasminogen activator and type 1 plasminogen activator inhibitor, and persistent fibrin deposition in chronic renal allograft failure.

Persistent fibrin deposition has been observed in kidneys undergoing chronic rejection, and has been suggested to contribute to the obliteration of the vasculature in these grafts. The mechanisms leading to it are not clear. Fibrinolysis, the process to remove fibrin in tissues, is initiated by tissue type plasminogen activator (tPA) and suppressed by type 1 plasminogen activator inhibitor (PAI-1). To investigate their roles in chronic rejection and fibrin deposition, we serially examined the expression of tPA and PAI-1 in an unmodified chronic rejection model, using a Fisher 344 to Lewis rat renal transplant, at 0, 2, 4, 6, 10, 12, 16 and 20 weeks post-transplantation (N = 4 rats/time point in each group). We also analyzed fibrin deposition and the development of chronic changes in the grafts. Our results show that tPA was up-regulated only in the acute phase of rejection (P < 0.05), whereas PAI-1 was induced and persistently expressed during the progressive phase of chronic rejection, together with persistent fibrin deposition in the grafts. Immunohistochemistry showed PAI-1 was mainly localized to the damaged/proliferative vascular intima. The results suggest that persistent induction of PAI-1 may be responsible for the continuance of fibrin deposition, which is associated with irreversible damage and chronic graft loss.

Animals↗
PubMed1996

Up-regulation of type 1 plasminogen activator inhibitor messenger RNA with thrombotic changes in renal grafts.

Small vessel thrombosis is a prominent feature in kidneys undergoing vascular rejection. Type I and type 2 plasminogen activator inhibitors (PAI-1 and PAI-2, respectively) are known to mediate thrombosis. To examine the potential role of PAI-1 and PAI-2 in the mediation of vascular injury, the relationship and the time course of gene expression of PAI-1 and PAI-2 with the thrombotic changes in renal grafts were investigated in an unmodified rejection model in rats. Orthotopic renal transplantation was performed from Lewis to dark agouti (DA) rats and from DA to DA isografts; untreated normal rat kidneys were used as controls. The rats were killed on days 1-9 posttransplantation (n=18 in each allograft and isograft group). The grafts were analyzed by histopathology, in situ mRNA hybridization and Northern blot methods. The results show that PAM mRNA was first detected at day 4, when the thrombotic changes in the grafts were first seen, and that this relationship persisted during the time course observed to day 9. There was no detectable PAI-1 mRNA in the control groups and no PAI-2 in either group. In situ hybridization showed that PAI-1 positive cells were predominantly located in the cortical interstitium, consistent with the distribution of interstitial microthrombi. These results provide experimental evidence that the thrombotic changes in rejecting allografts are associated with the up-regulation of PAI-1 in the donor tissue, whereas PAI-2, from our results, does not seem to influence these changes. The data are consistent with a role for PAI-1 in the pathogenesis of vascular rejection.

Animals↗
PubMed1996

Allograft immune response with sCR1 intervention.

The deposition of complement (C) components on tissues of transplanted organs may induce many proinflammatory responses. The role of such C activation in allograft rejection is uncertain. We addressed this question by inhibiting C at the level of the C3 and C5 convertases, preventing C activation and progression of its cascade, using recombinant human soluble complement receptor 1 (sCR1) in an unsensitized rat renal allograft model. Fully MHC disparate Lewis to DA rat renal allograft recipients given 25 mg/kg sCR1 daily, with saline-treated allograft recipients as controls (n = 15 in each group), were sacrificed from day 1 to day 5 post-transplant, and examined histopathologically, and for the deposition of C3 and C5b-9 membrane attack complex (MAC), and for the presence of leucocyte antigen markers. Treated animals demonstrated a reduction in vascular injury and cellular infiltration, coincident with reduced C deposition. Flow cytometric analysis of leucocyte subpopulations in the spleen showed a reduction in activated (CD25 positive) B and T cells in treated animals, compared to saline treated controls. The results suggest that C inhibition with sCR1, in an unsensitized model of allograft rejection, was able to suppress the vascular and cell mediated components of tissue injury. The data support not only a role for C in antibody and possibly cell mediated cytotoxicity in the graft, but also suggest a role in the primary immune response leading to both T cell and B cell activation.

Animals↗
PubMed1996

Effects of complement inhibition with soluble complement receptor-1 on vascular injury and inflammation during renal allograft rejection in the rat.

Complement is both an effector of the humoral immune response and a stimulator of leukocyte activation. To examine the influence of complement on the allograft response, we inhibited complement using recombinant human soluble complement receptor-1 (sCR1; TP10), in an unsensitized model of rat renal allograft rejection. Lewis to DA renal transplant recipients were treated daily with 25 mg/kg sCR1 or saline and sacrificed on days 1 to 5 after transplant. Transplanted organs were examined histologically and immunohistochemically for leukocyte subset markers and for the third component of complement, C3, and membrane attack complex deposition. A second set of recipients was followed from day 5 to day 9 to assess graft survival. sCR1-treated recipients displayed > 90% inhibition of plasma complement activity and a marked reduction in tissue C3 and membrane attack complex deposition. Inactivation of complement reduced the vascular injury such that there was almost complete sparing of vascular damage in day 5 sCR1-treated rats. There was a significant reduction in infiltrating leukocytes by day 5 after transplant, and complement inhibition delayed the time to reach a histologically defined end point of graft survival from 5 days in controls to 9 days in the sCR1-treated group. These results imply that the vascular and cell-mediated injury arises, in part, from complement activation. The partial inhibition of these injuries by sCR1 may have functional implications for strategies to inhibit allograft rejection.

Acute Disease↗
PubMed1994

Evidence of alloreactive T suppressor cells in the maintenance phase of spontaneous tolerance after orthotopic liver transplantation in the rat.

In the DA-->PVG rat strain combination liver allografts are spontaneously tolerated and induce a state of systemic tolerance towards donor antigens. Thirty long-term (> 100 days)-surviving liver recipients in this strain combination were used to study humoral and cellular mechanisms of tolerance in the maintenance phase after liver transplantation. In the popliteal lymph node assay splenocytes of liver grafted or native PVG rats displayed a similar proliferative response in (DAxPVG)F1 hybrid rats, indicating an unaffected capacity of these cells to mount graft-versus-host reactions. Using adoptive transfer assays T-splenocytes (1x10(7) cells, MRC Ox 19+, MRC Ox12-, CD 5) of long-term-surviving liver recipients were able to suppress the rejection of DA hearts (donor-specific), of DA and AO kidney grafts in irradiated (300 cGy) PVG recipients. Serum of long-term-surviving liver recipients was able to suppress the rejection of semiallogeneic (DAxPVG)F1 heart grafts after double injection of 1 ml serum, while fully allogeneic DA heart as well as skin and pancreaticoduodenal grafts were resistant to passive enhancement. The results confirm the development of spontaneous tolerance after liver grafting in the DA-->PVG strain combination, and the demonstration of T cells with suppressive capacity in adoptive transfer experiments indicates a role for T suppressor cells in the maintenance phase of spontaneous tolerance after liver grafting.

Animals↗
PubMed1994

Use of fluorochromes for direct enumeration of total bacteria in environmental samples: past and present.

Understanding the role of bacteria in microbial food webs is intimately connected to the methods applied in the direct enumeration of bacteria. We have examined over 220 papers describing studies in which fluorochrome staining followed by epifluorescent microscopic direct counts was used to estimate total bacterial abundances. In this review, we summarize patterns in the use of 3,6-bis[dimethylamino]acridinium chloride (acridine orange) and 4',6-diamidino-2-phenylindole (DAPI), the two stains most frequently used in bacterial enumeration. The staining of samples with these fluorochromes, followed by filtration and direct counting of bacterial cells on filter surfaces, has become routine over the past 10 years. We examine trends in features of the standard direct count methods, such as sample preservation and preparation techniques, membrane filter types used, applied stain concentrations, duration of staining, and counting strategies, in relation to the types of samples being examined. The high variability in bacterial counts observed within similar sample types may be partially accounted for by differences in methods. Synthesizing review findings, we include a recommended method for the direct enumeration of bacteria in environmental samples.

Bacteria↗
PubMed1994

A study of home care quality management data-collection systems.

A recent study reveals that home care agencies have inconsistent standards for quality management and inefficient data-collection systems. Are currently available data-collection systems suited for home care needs, or should they be redesigned?

Data Collection↗
PubMed1993

Analysis of activated T cell infiltrates in rat renal allografts by gamma camera imaging after injection of 123iodine-interleukin 2.

Activated T cells bearing receptors for interleukin 2 (IL-2) play an important role in immunity and in immunopathological processes such as allograft rejection. In order to investigate the presence of activated T cells in lymphocytic infiltrates in transplanted kidneys, we investigated the uptake and retention of radioactivity after an intravenous injection of radioiodinated IL-2 in experimentally transplanted rats. IL-2 was enzyme radiolabelled with 123iodine using a glucose oxidase/lactoperoxidase method and shown to retain specific binding on an IL-2 receptor positive cell line, C58E6. To examine the kinetics of 123iodine-interleukin 2 (123I-IL-2) uptake in vivo, animals that had been transplanted five days previously with allogeneic or syngeneic grafts were injected with 123I-IL-2 and then imaged using an external gamma camera. Radioactivity was measured at time points up to 240 min after intravenous injection of 123I-IL-2. Four groups of animals were examined: allogeneic grafts (n = 7); syngeneic grafts (n = 6); ischaemic native kidneys (n = 5) all following injection with 123I-IL-2; and allogeneic transplants (n = 5) after injection of 123I-lactalbumin, an irrelevant molecule of similar molecular weight to IL-2. The peak radioactivity after injection was measured and the amount of radioactivity retained in the graft at increasing time intervals after injection was expressed as a function of initial peak radioactivity. At four hours after injection of 123I-IL-2, mean retention of activity by rejecting grafts was 77(+/- 2.68)% of peak activity, compared to 45(+/- 6.38)% in syngeneic controls (p < 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗