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Biomedical subjects

M W Flye

Publications and source records attributed to M W Flye.

At least 55 records · Page 3Linked to original sources
PubMed1996

Gadolinium chloride inhibits Kupffer cell nitric oxide synthase (iNOS) induction.

Kupffer cells (KC) are the phagocytic macrophages of the liver. The rare earth metal, gadolinium (GdCl3), is a lanthanide, which, after phagocytosis by the KC, has been found to alter various aspects of KC physiology. In this study, we describe for the first time that the in vivo administration of GdCl3 to rats decreases the release of NO by isolated rat KC in response to lipopolysaccharide. Western blot analysis shows decreased expression of both inducible nitric oxide synthase as well as total cellular calmodulin after GdCl3 treatment. Possible mechanisms for this phenomenon are suggested.

Animals↗
PubMed1996

Inhibition of phosphatidylinositol-3'-kinase prevents induction of endotoxin tolerance in vitro.

Previous studies have shown an increase in the expression of phosphatidylinositol-3'-kinase (PI-3'-K) in macrophages from endotoxin tolerant (ET) rats. This implicates PI-3'-K cell signaling in attenuated macrophage responsiveness to lipopolysaccharide (LPS). These experiments examined the effects of selective pharmacologic inhibition of PI-3'-K in an in vitro model of ET. To induce ET, RAW 264.7 macrophages cultured in RPMI 1640 with 10% fetal calf serum were initially exposed to 10 ng/ml LPS (E. coli 0111:B4) for 19 hr. Non-tolerant cells received an equal volume of phosphate buffered saline. Some cultures were also incubated with the specific PI-3'-K inhibitor wortmannin (10 nM) during this tolerizing period. Cells were then washed and re-challenged with 100 ng/ml LPS for 24 hr. Next, macrophage tumor necrosis factor-alpha (TNF-alpha) and nitrite production were measured as indicators of ET induction. Macrophage TNF-alpha production decreased significantly while nitrite production increased significantly following ET induction. Specific inhibition of PI-3'-K prevented this decrease in TNF-alpha and increase in nitrite production in ET macrophages. Production of each mediator returned to levels not different than in non-tolerant macrophages. In this in vitro model of macrophage ET, pharmacologic inhibition of the PI-3'-K signaling pathway prevented the induction of LPS tolerance as measured by the production of two inflammatory mediators.

Androstadienes↗
PubMed1996

Femoropopliteal bypass for claudication: vein vs. PTFE.

The vascular graft of choice for femoropopliteal bypass in patients with intolerable claudication is controversial. We retrospectively reviewed our experience with 239 patients suffering from claudication secondary to superficial femoral artery obstruction. Femoropopliteal reconstruction was performed with saphenous vein to the below-knee popliteal artery in 66 patients (BK-vein). Polytetrafluoroethylene (PTFE) was used in 128 patients as a bypass graft to the above-knee popliteal artery (AK-PTFE) and 45 patients had a PTFE graft to the below-knee popliteal artery (BK-PTFE). All patients were enrolled in a postoperative graft surveillance program with graft revision when appropriate. There was one perioperative death (0.4%). Primary patency at 5 years for AK-PTFE, BK-PTFE, and BK-vein was 58.0%, and 60.3%, respectively, and was not significantly different among the graft groups. Graft revision for failed/failing grafts resulted in 5-year secondary patency rates of 79.2% (AK-PTFE), 73.3% (BK-PTFE), and 74.4% (BK-vein). These secondary patency rates were not statistically different. Eventual conversion to a vein graft in patients initially treated with PTFE maximized patency in the femoropopliteal segment with 5-year patency rates of 84.6% and 93.0% for the AK-PTFE and BK-PTFE graft groups, respectively. Major leg amputation was necessary during the entire course of the study in eight (3.3%) patients. We conclude that long-term patency rates for femoropopliteal bypass in patients with intolerable claudication are similar for PTFE and autologous saphenous vein grafts.

Aged↗
PubMed1996

Revascularization of the solitary kidney: a challenging problem in a high risk population.

BACKGROUND: Patients with significant atherosclerotic stenosis involving the artery to a solitary functioning kidney present a clinical challenge. METHODS: From August 1987 through August 1995, 35 of these patients (average age, 68.4 +/- 6.9 years) were treated. Comorbid conditions included previous myocardial infarction in 23% of the patients, congestive heart failure (CHF) in 34%, chronic obstructive pulmonary disease in 20%, and diabetes in 20%. The average creatinine level of the patients was 2.5 +/- 1.5 mg/dl. Indications for revascularization were hypertension in 86%, hypertensive crisis with CHF in 17%, and renal insufficiency in 69%. Procedures performed included 19 extra-anatomic bypasses, 8 concomitant with infrarenal aortic reconstruction and 2 concomitant with thoracoabdominal aortic aneurysm repair; 1 visceral segment endarterectomy; 1 renal artery endarterectomy with reimplantation; I superior mesenteric to renal artery bypass; 1 aortorenal bypass; and 2 percutaneous angioplasties with staged nephrectomies. RESULTS: At discharge, 91% of patients had stable or improved renal function with an average creatinine level of 1.7 +/- 0.8 mg/dl. Hypertension was cured or improved in 85%. Perioperative mortality was 6%, and major morbidity was 43%, including the need for permanent (9%) and temporary (9%) dialysis, respiratory insufficiency (18%), two early reoperations, six cardiac complications, one case of gastrointestinal bleeding, and one stroke. In the follow-up period (mean duration, 39.2 months), survival has been 73%, and no additional patients have required dialysis. CONCLUSIONS: Although significant perioperative morbidity exists in this high risk population, the long-term preservation of renal function and improvement in hypertension make solitary renal revascularization worthwhile.

Aged↗
PubMed1996

Long-term survival of rat to mouse cardiac xenografts with prolonged blockade of CD28-B7 interaction combined with peritransplant T-cell depletion.

BACKGROUND: The hCTLA4Ig/mCTLA4Ig fusion protein of the extracellular domain of human/mouse CTLA4 and the Fc portion of the human/mouse immunoglobulin G1 block the CD28/B7 costimulatory T-cell activation pathway. We evaluated the effect of prolonged B7-CD28 blockade, T-cell depletion, or both on rat to mouse cardiac xenografts. METHODS: C57BL/6 (H-2b) mice receiving infant Wistar Furth (RT1u) rat cardiac xenografts were treated with anti-CD4 (GK1.5) and anti-CD8 (2.43) monoclonal antibodies (mAb; 0.2 mg intravenous each) on days -2 and 0, hCTLA4Ig or mCTLA4Ig every other day from day 0 until day 14 and then twice a week until day 50 or day 100, or both. Changes in cellular reactivity were assayed by mixed lymphocyte culture and cell-mediated cytotoxicity and the development of cytotoxic antibodies was serially measured after transplantation. RESULTS: Either human CTLA4Ig or murine CTLA4Ig alone led to significant prolongation of rat to mouse cardiac xenografts (median survival time [MST], 22 or 26 days, respectively [p = 0.008], versus control). hCTLA4Ig given for 50 days in combination with two doses of anti-CD4/CD8 monoclonal antibodies further prolonged graft survival (MST, 61 days; p versus control < 0.0001). In this combination, when hCTLA4Ig was continued until day 100, the graft survival was further prolonged (MST, 119 days). mCTLA4Ig for 100 days plus anti-CD4/CD8 similarly prolonged rat xenograft survival (MST, 94 days). However, all cardiac xenografts eventually failed, primarily from humoral rejection. Cytotoxic antibody titers rose rapidly only in animals rejecting a graft, and suppressed cell-mediated immunity had completely recovered in rejecting recipients. CONCLUSIONS: Blockage of the CD28-B7 costimulatory interaction can inhibit both humoral and cell-mediated immune responses and result in the prolonged acceptance of rat to mouse cardiac xenografts. Longer administration of CTLA4Ig and anti-CD4/CD8 monoclonal antibodies further prolongs but does not achieve indefinite survival of rat cardiac xenografts.

Abatacept↗
PubMed1996

Induction of allogeneic islet survival by intrahepatic islet preimmunization and transient immunosuppression.

Induction of tolerance to fully major histocompatibility complex (MHC)-mismatched rat islet allografts implanted at two different islet transplant sites (liver and kidney capsule [KC]) was examined. Streptozotocin-induced diabetic Lewis (RT1(1)) rats remained hyperglycemic (> 200 mg/dl) after intrahepatic preimmunization by injection of 200 low-temperature cultured (24 degrees C for 7 days) Wistar-Furth (WF, RT1u) rat islets into the portal vein with one injection (1 ml) of rat antilymphocyte serum intraperitoneally. Three weeks later, 1,200 WF islets that had been cultured to remove passenger lymphoid cells were transplanted into the liver via the portal vein or under the KC. The intrahepatic transplants survived 60.2 +/- 11.9 days, and all six of the KC transplants maintained normoglycemia for > 100 days after the preimmunization regimen. In contrast, survival of fresh islet transplants was not significantly improved by this preimmunization protocol at either transplantation site. This study demonstrates that indefinite islet allograft survival can be achieved across a full MHC mismatch by intrahepatic preimmunization with a small number of cultured donor islets and a brief period of immunosuppression followed by transplantation of low-temperature cultured donor islets.

Animals↗
PubMed1996

Successful treatment of intraoperative malignant hyperthermia during renal transplantation.

Malignant hyperthermia is a complication of general anesthesia that is especially problematic if it occurs during renal transplantation because myoglobinemia, shock, and ischemia play a role in injuring the transplanted kidney. In this report, we describe a case of malignant hyperthermia, its clinical course, and the measures taken to successfully treat it and preserve the function of a kidney allograft.

Adult↗
PubMed1996

Serum bilirubin pigments covalently linked to albumin.

The serum levels of biliprotein (i.e., the bilirubin fraction covalently linked to albumin) were measured in patients with hepatobiliary disease by two analytical procedures: reverse-phase anion-exchange chromatography (Bond-Elut) and anion-exchange chromatography. Bond-Elut extracts contained tetrapyrroles, allowing the analysis of biliprotein, bilirubin, and bilirubin conjugates, whereas the anion-exchange method converted tetrapyrroles to azodipyrroles (protein-free), which were adsorbed on the resin, with the exception of azopyrrole of biliprotein (covalently linked to albumin), which passed through the resin and could be analyzed. In contrast, other cationic polymer columns did not completely bind the alkaline diazo derivatives of unconjugated bilirubin, conjugated bilirubin, or the azopyrrole of biliprotein not bound to albumin. Therefore, a comparison was made of these two methods using the original anion-exchange resin. Both procedures gave similar values for serum levels of biliprotein in patients with high levels of total bilirubin.

Bile Pigments↗
PubMed1995

Donor-specific transfusions have long-term beneficial effects for human renal allografts.

From April, 1980 to November, 1988, 163 one or two haplotype-mismatch living related (LR) or living unrelated (LUR) potential renal transplant recipients received three 200 ml aliquots of donor specific transfusion (DST) at biweekly intervals with concomitant azathioprine (2 mg/kg/day). Following transplantation, only prednisone and azathioprine were given for immunosuppression. The results for the DST group are compared with those for HLA identical living recipients (57 patients) transplanted during this same interval (1980-1988). Comparison is also made with a group of one or two haplotype-mismatched living donor recipients (54 patients) not treated with DST but with triple drug therapy (prednisone, azathioprine, cyclosporine) and antithymocyte globulin (ATG) or OKT-3 induction. Permanent T cell crossmatch sensitization occurred in 11 of 163 patients (7%). Successful DST donor transplants were performed between 121 one HLA haplotype-mismatched, 14 two HLA haplotype-mismatched LR, and 7 two haplotype-mismatched LUR pairs. Actual one- and five-year graft survivals were 94%, 100%, 100%, and 72%, 85%, and 71%, respectively, for these three subgroups of DST treated patients. The graft survival for all DST pretreated recipients at one, five, and ten years was comparable to the HLA-identical group (94%, 79%, 64% vs. 91%, 80% and 77%). At a mean follow-up of 10 1/2 years, 54% (80 patients) of the entire group of 147 patients transplanted after DST have functioning transplants with a mean serum creatinine of 1.7 mg/dl. Fifteen percent of DST patients (21 patients) died with a functioning graft 2 to 132 months after transplantation, 26% (37 patients) rejected the DST graft after 1 to 128 months, and 6% (9 patients) were lost for nonimmunological reasons. No lymphoproliferative disease developed in the DST group and the incidence of cytomegalovirus sepsis was only 2% (3 patients). The long-term beneficial effects of DST on renal allograft survival and function and the lower incidence of the complications of nonspecific immunosuppression should encourage increased utilization of DST in renal transplantation.

Azathioprine↗
PubMed1995

Ras-transduced diethylnitrosamine-treated hepatocytes develop into cancers of mixed phenotype in vivo.

The cell of origin of hepatocellular carcinoma (HCC) is controversial. A method for marking cells of different lineages in vivo and then determining their carcinogenic potential should resolve this issue. A retroviral vector expressing activated ras and beta-gal genes (Ras-gal) was transferred into adult rat hepatocytes in vivo, and some animals were treated with diethylnitrosamine (DEN). Bile ductule cells and the putative stem cells of the liver (the oval cells) did not appear to be transduced by this method. At 1 month after transfer, 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside staining was performed on transduced rat livers to determine the blue cluster size. Eight % of the clusters in Ras-gal-transduced, DEN-treated livers contained at least twice as many cells as the largest cluster in Ras-gal-transduced, DEN-untreated rats, demonstrating that they had acquired markedly abnormal growth properties. When the retroviral vector containing beta-gal without ras (Gal-509) was transferred into DEN-treated rats, 2.5% of the cells were present in clusters containing at least twice as many cells as the largest cluster in Gal-509-transduced, DEN-untreated animals. Thus, p21-ras may increase the percentage of cells that acquire mutations in response to DEN, or it may behave synergistically with other mutations to increase the replication rate of cells. Occasional foci in Ras-gal-transduced, DEN-treated rats had extramedullary hematopoiesis. Forty % of the Ras-gal-transduced, DEN-treated rats developed unifocal HCC, mixed HCC/cholangiocarcinoma (CC), or CC at 3-6 months after transduction, suggesting that hepatocytes can develop into HCC or CC if sufficient genetic alterations occur.

Animals↗
PubMed1995

Endotoxin tolerance alters phospholipase C-gamma 1 and phosphatidylinositol-3'-kinase expression in peritoneal macrophages.

Although the molecular signaling mechanisms underlying macrophage endotoxin (LPS) responsiveness are not fully understood, alterations in intracellular phosphatidylinositol (PI) metabolism appear to contribute. We evaluated the effects of endotoxin tolerance (ET) induction upon peritoneal macrophage (PM) expression of the principal PI enzymes phospholipase PLC-gamma 1 (PLC-gamma 1) and phosphatidylinositol 3'-kinase (PI-3-K). Rats received either 5 mg/kg LPS (ET) or phosphate-buffered saline (nontolerant, NT) which enabled 88% of ET and 25% of NT to survive a 25 mg/kg LPS dose 3 days later. PM were harvested by lavage on Day 3 from both ET and NT rats. Following overnight culture, 5 x 10(6) PM in serum-free media were stimulated with 5 ng/ml LPS for 0 to 30 min. Cell lysates fractionated by SDS-PAGE were transferred to nitro-cellulose and blotted with PLC-gamma 1, PI-3'-K, and phosphotyrosine (4G10) monoclonal antibodies. Western immunoblots were developed by enhanced chemiluminescence and quantitated by densitometry. Unlike NT cells in which PLC-gamma 1 was expressed constitutively and increased with LPS stimulation, PLC-gamma 1 expression in ET cells stimulated with LPS was markedly reduced in three separate experiments. In contrast, ET cells expressed considerably higher concentrations of PI-3'-K to NT cells. Patterns of protein tyrosine phosphorylation were similar in both NT and ET cells regardless of LPS stimulation. The development of endotoxin tolerance decreased PLC-gamma 1 expression and markedly amplified PI-3'-K expression in macrophages. PI-3'-K-generated second messengers may contribute to unique signaling pathways responsible for tempered cellular responses to LPS.

Animals↗
PubMed1995

Cytolytic T lymphocytes from human renal allograft biopsies are tissue specific.

The cytolytic activity of T lymphocytes infiltrating renal allografts from recipients undergoing episodes of acute cellular rejection was studied. These T-cell populations, composed of both CD4+ and CD8+ cells, demonstrated significant cytolytic activity against both donor-derived KCLs and B-LCLs. In five of 21 biopsy-derived lines greater cytolytic activity was measured against donor KCLs compared to donor B-LCLs, suggesting the presence of kidney antigen-specific, MHC-restricted clones. Clones developed by stimulation with donor B-LCLs lysed both donor B-LCLs and KCLs while clones developed on donor KCLs as stimulator cells showed tissue specificity. Three of 26 clones recognized tissue-specific antigens in the context of donor MHC class I antigens lysing donor KCLs but not B-LCLs. These data demonstrate that a subpopulation of T cells recognizing kidney-specific antigens are present in biopsies of renal allograft recipients undergoing acute cellular rejection. This subpopulation of tissue-specific cytotoxic T lymphocytes may prove to contribute significantly to the pathology of allograft rejection.

Biopsy↗
PubMed1995

Donor-specific transfusion and donor bone marrow infusion in renal transplantation tolerance: a review of efficacy and mechanisms.

The ultimate goal in transplantation is modulation of the immune response to produce tolerance without immunosuppression. To date only a state of pseudotolerance for the allograft has been achieved through the use of potent pharmacologic and biologic manipulations. Despite these manipulations to prevent acute rejection, chronic rejection eventually results in graft failure. Thus, different strategies have been sought to induce tolerance and prevent acute and chronic rejection. Historically, donor-specific transfusion (DST) was one such strategy attempted. Donor-specific transfusion has been used primarily in living donor organ transplantation. With the concern that DST may sensitize patients, thereby preventing transplantation and introduction of cyclosporine, the use of DST was curtailed. More recently, donor bone marrow (DBM) infusion at the time of cadaveric transplantation has been used to facilitate development of microchimerism and tolerance to abrogate acute and chronic rejection. However, DBM infusion may predispose the recipient to graft-versus-host disease and is not easily accomplished in living donor organ transplantation. The potentially immunomodulating and tolerizing mechanisms of DST and DBM infusion are similar and include induction of anergy, stimulation of anti-anti-HLA antibodies, provision of soluble HLA antigen, suppressor cell and/or veto cell activity, clonal deletion, regulation of cell surface molecules, regulation of cytokines, promotion of microchimerism, or a combination of these. Of these mechanisms, microchimerism with the concomitant persistence of soluble donor HLA antigen is felt by many to be the most important. Although microchimerism is detectable in many patients who are tolerant of their grafts, there is no clear evidence that chimerism is responsible for the induction or maintenance of tolerance.

Animals↗