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Biomedical subjects

M W Flye

Publications and source records attributed to M W Flye.

At least 91 records · Page 5Linked to original sources
PubMed1994

Autoregulation by eicosanoids of human Kupffer cell secretory products. A study of interleukin-1, interleukin-6, tumor necrosis factor-alpha, transforming growth factor-beta, and nitric oxide.

OBJECTIVE: Methods employed previously to analyze the secretory behavior of rodent Kupffer cells (KC) were used to examine the human KC's secretory response to lipopolysaccharide (LPS). SUMMARY BACKGROUND DATA: As the resident hepatic macrophage, the KC resides at the interface between the portal and systemic circulations. Consequently, this cell may play an integral role in the immune response to antigens and bacteria in the sinusoid. Study of cytokine production by the KC has relied predominantly on the rat as the source of these cells. Whether human KCs respond similarly to rat KCs after LPS stimulation has been a matter of speculation. METHODS: Kupffer cells obtained from seven human livers were tested under conditions identical to those used to study rat KCs. Kupffer cells rested for 12 hours after isolation were stimulated with LPS (2.5 micrograms/mL). Arginine concentration in the culture medium varied from 0.01 to 1.2 mM. To examine the role of eicosanoids, parallel culture wells received indomethacin (10 microM). Culture supernatants were assayed for interleukin-1 (IL-1), interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), transforming growth factor-beta (TGF-beta), prostaglandin E2 (PGE2), and nitric oxide. RESULTS: Similar to the rat KC, LPS-stimulated human KCs released IL-1, IL-6, TNF-alpha, TGF-beta, and PGE2. However, unlike rat KCs, nitric oxide could not be detected, regardless of whether the human KCs were exposed to LPS, interferon-gamma (INF-gamma), or LPS + IFN-gamma. Similar to rat KCs, indomethacin prevented PGE2 release while significantly upregulating TNF-alpha, IL-1, and IL-6, but not TGF-beta, consistent with an autoregulatory control of eicosanoids over proinflammatory cytokines. As has been shown in the rat, physiologic levels of L-arginine (0.01 mM) significantly enhanced LPS-induced PGE2 secretion relative to the response in medium containing standard L-arginine concentration (1.2 mM); however, unlike the rat KC, the human's cytokine response to LPS was not downregulated by this enhanced PGE2 release. CONCLUSIONS: Although many functional features are shared by rat and human KCs, significant differences do exist. Such discrepancies reinforce the need to proceed with caution when generalizing from the results obtained in other species to human physiology.

Adult↗
PubMed1994

Induction of immune unresponsiveness to concordant islet xenografts by intrahepatic preimmunization and transient immunosuppression.

Streptozotocin-induced, diabetic mice (C57BL/6) were preimmunized by injecting 25 low temperature, cultured Wistar-Furth (WF) rat islets into the portal vein, and the recipients received one injection of mouse and rat antilymphocyte sera. 3 wk later, fresh WF islets were transplanted under the kidney capsule of the preimmunized recipients, and normoglycemia was maintained in all 13 recipients for 60 d. Removal of the grafts at 60 d returned the mice to a diabetic state. Transplants of fresh WF islets under the kidney capsule without pretreatment of the recipients had a mean survival time of 16.5 +/- 2.5 d. These findings demonstrate that immune unresponsiveness can be achieved across a concordant, islet xenograft barrier within 3 wk after intrahepatic preimmunization with a small number of donor rat islets and transient immunosuppression with antilymphocyte sera.

Animals↗
PubMed1994

Induction of tolerance to islet xenografts in a concordant rat-to-mouse model.

Induction of tolerance to concordant rat islet xenografts (150 Wistar-Furth [WF] islets) in streptozocin-induced (STZ) diabetic mice (C57BL/6) was determined at three different sites for islet implantation (thymus, kidney capsule, and liver). Islets transplanted into the thymus or kidney capsule were either fresh or cultured at 24 degrees C for 7 days, and the mice received a single injection of either anti-mouse lymphocyte serum (MALS) alone or anti-rat lymphocyte serum (RALS) and MALS. Islets transplanted into the liver via the portal vein were cultured at 24 degrees C for 7 days, and the mice received a single injection of MALS and RALS. To document the induction of tolerance, recipients with islet xenografts surviving > 100 days were made diabetic again by STZ (thymus and liver) or nephrectomy (kidney capsule) and received a second transplant of 150 fresh WF islets in the kidney capsule. Kidney capsule placement of fresh or cultured islets with MALS alone or MALS and RALS did not induce tolerance in a significant number of recipients. The intrathymic transplantation of fresh or cultured islets with MALS alone resulted in prolonged WF islet xenograft survival (mean survival time of 39.7 +/- 7.9 days) but did not result in tolerance, whereas the administration of MALS and RALS with the intrathymic placement of fresh or cultured islets induced tolerance in approximately 50% of the mice. Intrahepatic transplantation of cultured islets with MALS and RALS resulted in tolerance to donor islets in 90% of the recipients. Donor specificity was evaluated by a third major histocompatibility complex-disparate fresh Lewis islet xenograft.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

The Kupffer cell in endotoxin tolerance: mechanisms of protection against lethal endotoxemia.

Kupffer cells (KC) of the hepatic sinusoid respond to endotoxemia by producing mediators which promote or inhibit systemic inflammatory responses. Sublethal lipopolysaccharide (LPS) pretreatment confers tolerance to the lethality of a subsequent LPS exposure. However, the precise role of the KC in endotoxin tolerance (ET) remains unclear. This study evaluated the effect of ET induction upon the rat KC production of the mediators tumor necrosis factor-alpha (TNF-alpha), prostaglandin E2 (PGE2), and interleukin-6 (IL-6), and upon the in vivo phagocytic capacity of the KCs. 3 days prior to KC isolation, age-matched rats received either 5 mg/kg LPS (ET) or normal saline (nontolerant, NT), which protected 100% of the ET rats against an LPS dose 3 days later which was lethal in 72% of NT rats. On an in vitro LPS rechallenge, ET KC produced significantly lower amounts of TNF than NT KC (p < .01). In contrast, the ET KC produced significantly more PGE2 (p < .05) and IL-6 (p < .001) than the NT KC. The percentage of KC phagocytosing fluorescent latex spheres in vivo was increased 7-fold in the ET rats. Thus, ET induction, which protects rats against subsequent lethal endotoxemia, selectively alters KC mediator production and phagocytic capacity. These findings strongly implicate the KC in the mediation of early endotoxin tolerance.

Animals↗
PubMed1994

Portal vein administration of donor cells promotes peripheral allospecific hyporesponsiveness and graft tolerance.

BACKGROUND: Major histocompatibility complex-mismatched Lewis rate cardiac grafts (inferior vena cava drained) are promptly rejected by Buffalo recipients (mean survival time, 7 days). Ultraviolet-B (UVB) irradiation modulates donor immunogenicity and down-regulates in vitro immune responses. We found that Buffalo recipients (BUF; RT1b) preimmunized with 2.5 x 10(7) nontreated Lewis spleen cells (LEW; RT1(1)) by portal venous (PV) injection 7 days before a portal vein-drained heterotopic graft of LEW heart resulted in only 20% surviving long-term (n = 10) (naive rats rejected a cardiac allograft drained into the portal vein in mean survival time of 13.5 days [n = 5]). In contrast, BUF recipients receiving PV injection of UVB-treated LEW spleen cells permitted 70.6% indefinite allograft survival (n = 34) with a mean survival time of greater than 150 days. METHODS: These studies examined the effect of untreated or UVB-irradiated donor spleen cells administered intraportally on the recipient's cell-mediated responsiveness as serially measured by the mixed lymphocyte culture and limiting dilution analysis of T-helper precursors (pTH) and cytotoxic precursors (pCTL). Survival of heterotopic cardiac allografts transplanted with PV drainage at various intervals after PV alloantigen administration were correlated with changes in these in vitro studies. RESULTS: The precursor frequencies in the spleens of BUF recipients given nontreated LEW spleen cells intraportally 7 days previously were 1/16,170 (pTH) and 1/11,929 (pCTL), whereas those in the spleen of BUF recipients receiving UVB-treated LEW spleen cells decreased to 1/152,409 pTH and < 1/5 X 10(5) pCTL. These hyporesponsive BUF spleen cells responded normally to the third-party alloantigen, irradiated ACI spleen cells (RT1 alpha), with mixed lymphocyte culture tritiated incorporation of 133,220 cpm (control, 123,276 cpm), 1/13,364 pTH (control, 1/13,541), and 1/71,156 pCTL (control, 1/73,985), indicating that the hyporesponsiveness induced by UVB-treated donor spleen cells is antigen specific. The recovery of in vitro responsiveness correlates with decreasing cardiac graft survival to normal rejection times by 200 days after portal vein antigen administration. When 100 units/ml exogenous recombinant interleukin-2 (rIL-2), but not rIL-1 or rIL-4, was added to cultures, the hyporesponsiveness of spleen cells recovered to normal levels with tritiated incorporation of 201,227 cpm and pTH frequency of 1/32,227, suggesting that the hyporesponsiveness to donor antigen is due to anergized T-helper lymphocytes and not to clonal deletion. CONCLUSIONS: By administering properly modified alloantigen into the hepatic environment by portal vein inoculation, prolonged T-cell anergy can be induced, which allows the indefinite acceptance of donor-specific heterotopic cardiac allografts.

Animals↗
PubMed1993

Specific prolongation of allograft survival by a T-cell-receptor-derived peptide.

Allograft rejection results from the specific recognition by host CD8+ T cells of allogeneic major histocompatibility complex (MHC) molecules on the tissue graft. The specificity of this cellular response is determined by the molecular interaction of the T-cell receptor (TCR) on host T cells with the MHC molecule and its bound ligand on the grafted tissue. To better understand the precise manner by which the TCR interacts with the MHC-peptide complex and how to therapeutically intervene, we have studied the allogeneic response to the mouse class I MHC molecule Ld. In this report, the therapeutic potential of a synthetic peptide derived from the TCR V beta 8 variable region that predominates in responses to Ld was tested. This V beta 8-derived peptide was found to dramatically and specifically block the in vivo and in vitro allogeneic response to Ld. Furthermore, this specific blocking is not dependent upon the presence of V beta 8+ effector cells nor does the V beta 8 peptide bind to the Ld ligand binding cleft. We propose that this peptide functions as an antagonist, competing with the native TCR for recognition of the Ld molecule.

Amino Acid Sequence↗
PubMed1993

Major histocompatibility complex-specific prolongation of murine skin and cardiac allograft survival after in vivo depletion of V beta+ T cells.

The preferential usage of certain T cell receptor (TCR) V beta genes has been well established in several major histocompatibility complex (MHC)-restricted immune responses. However, V beta usage among allogeneic responses remains unclear. Because recent findings of ours and others indicate that V beta 8 predominates in certain Ld-restricted, peptide-specific responses, we examined the V beta 8 usage in allogeneic responses to Ld. To selectively recognize the Ld molecule, cells from BALB/c-H-2dm2 (dm2), the Ld-loss mutant mouse, were stimulated in vitro or in vivo with wild-type BALB/c cells. We report here that after the intraperitoneal administration of the anti-V beta 8 monoclonal antibody (mAb) F23.1, peripheral V beta 8 T cells were depleted from dm2 mice. This in vivo depletion abrogated the ability of dm2 splenocytes to mount a primary response to Ld molecules. This abrogation was specific, since the response of V beta 8-depleted dm2 cells to Kb/Db antigens was the same as that of control nondepleted dm2 cells. Furthermore, in vivo depletion of V beta 8 cells was found to cause a dramatic prolongation of Ld-disparate skin grafts (mean survival time [MST] 22.1 +/- 2.1 vs. 10.3 +/- 1.1 d for saline-treated controls, or 10.9 +/- 1.7 d for controls treated with mAb KJ23 to V beta 17). By contrast, V beta 8 depletion had no effect on recipients grafted with haplotype-mismatched skin or single Dk-locus-disparate skin. These findings demonstrate that V beta 8+ T cells predominate in allogeneic response to Ld but not other alloantigens. The effect of V beta 8 depletion was found to be even more dramatic on recipients grafted with Ld-disparate vascularized heart transplants (MST > 100 vs. 8.6 +/- 0.5 d for controls). In total, these findings establish the efficacy of using mAb to the V beta gene family to specifically and significantly enhance the survival of allografts. The implications of detecting V beta 8 usage in both alloreactive or MHC-restricted TCR responses to the same class I molecule are discussed.

Animals↗
PubMed1993

Clinical diagnostic applications of the polymerase chain reaction. Infection and rejection.

In this review, we describe the power and sensitivity of the polymerase chain reaction and indicate areas of clinical medicine in which it is currently being applied. As this technology and automation improves and is simplified, its realm of application will expand, but its major impact will continue to be its facilitation of early, specific diagnoses in infectious disease, genetics, cancer, and transplantation. Polymerase chain reaction is currently most useful in the care of opportunistic infections in immunosuppressed hosts and in making the distinction between infection and rejection in transplant recipients. As the technique becomes more widely available and less costly, its application should minimize the need for broad-spectrum antibiotic therapy of infections, lower costs, and perhaps even shorten hospitalization. These potential beneficial effects of polymerase chain reaction will be particularly relevant in the current cost-conscious health-care environment.

Anti-Bacterial Agents↗
PubMed1993

Lanthanide "blockade" of antigen-presenting cells suppresses lymphocyte proliferation by inducing nitric oxide synthesis.

Introduction of antigen into the portal venous circulation can induce a state of antigen-specific tolerance. Reversal of this phenomenon by the administration of gadolinium chloride (GD) before portal venous inoculation with antigen has been thought to result from the inhibition of macrophage phagocytosis by GD, thereby permitting passage of antigen through the hepatic sinusoids into the systemic circulation where sensitization can occur. We recently demonstrated that i.v. GD also greatly enhances the ability of the hepatic macrophage, or Kupffer cell (KC), to suppress lymphocyte proliferation in vitro. In this study, lymphocytes from ovalbumin (OVA)-immunized Lewis rats were cocultured with OVA plus KC or adherent splenocytes from syngeneic rats injected i.v. with GD or saline. Whereas small numbers of either hepatic or splenic untreated macrophages enhanced lymphocyte proliferation, both macrophage populations treated with GD were markedly inhibitory. Neither the presence of lymph node macrophages nor of normal KC was able to prevent the effect of GD-treated KC indicating a mechanism of active suppression rather than defective macrophage antigen uptake, processing, and presentation. While cyclooxygenase inhibition had no effect, the addition of inhibitors of nitric oxide synthase, i.e., aminoguanidine and NG-methyl-L-arginine, fully reversed the suppression of proliferation by GD-treated macrophages. Nitrite, but not PGE2, levels correlated with the degree of inhibition by the GD-treated macrophages. The suppression of lymphocyte proliferation and elevated supernatant nitrite levels associated with the presence of GD-treated macrophages were reversed when the macrophages were preincubated with EDTA indicating that GD's effect was directly on macrophages and due to its persistence in cultured cells. The findings of this study suggest that GD's in vivo prevention of portal venous tolerance may be due to enhanced nitric oxide synthesis whose effect on lymphocytes could be to inhibit adhesion or replication in the hepatic sinusoid.

Animals↗
PubMed1993

In vivo hepatocyte transduction with retrovirus during in-flow occlusion.

Gene therapy research would be facilitated by a technically simple procedure for transducing hepatocytes in vivo. Previously reported methods have employed partial hepatectomy followed 24 hr later by asanguineous perfusion of the regenerating liver with retrovirus. We have developed a simpler method of in vivo transduction in which we deliver an intraportal bolus of retrovirus to the regenerating rodent liver during a brief period of hepatic in-flow occlusion. On Day 0, adult male Sprague-Dawley rats (N = 19) underwent 70% hepatectomy to induce hepatocyte replication. On Day 1, retroviral supernatant was harvested from an amphotropic retroviral packaging cell line that packaged an LNL6-derived vector containing the cytomegalovirus promoter driving expression of the Escherichia coli beta-galactosidase (beta gal) gene. Twenty-four hours after partial hepatectomy, experimental rats (N = 17) received 6 x 10(5) colony-forming units of retrovirus by intraportal injection during a 3-min occlusion of the hepatic artery and portal vein. Control rats (N = 2) received intraportal medium (without retrovirus), also during in-flow occlusion. The procedure required 20-25 min, and the survival rate was 84%. Cryostat sections were prepared from liver biopsies obtained on Post-transduction Days 8 and 15 and stained with 5-bromo-4-chloro-3-indolyl- beta-D-galactopyranoside to detect beta gal expression. Light microscopic examination of Day 8 sections from surviving experimental rats (N = 14) revealed 0.10-1.00% blue (i.e., transduced) hepatocytes per low power field, while sections from control rats (N = 2) exhibited no blue cells. Day 15 sections from experimental rats revealed a somewhat lower frequency of hepatocytes expressing beta gal.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Prolongation of small bowel allografts after intrathymic injection of donor alloantigen and ALS.

The increasing success of clinical small bowel transplantation in recent years has been due largely to improved nonspecific immunosuppression of the recipient, which carries significant morbidity. The induction of donor-specific tolerance would eliminate the risk of long-term immunosuppression while ensuring graft function and survival. We have demonstrated that the intrathymic injection of donor splenocytes with the simultaneous intraperitoneal administration of rabbit anti-rat lymphocyte serum results in indefinite donor-specific cardiac allograft survival in > 85% of recipients. In this study, we further examined the effect of this tolerance induction protocol on the more immunogenic small bowel allograft. Male Buffalo (RT1b) rats were exposed to donor alloantigen by an intrathymic injection of 25 x 10(6) MHC mismatched unfractionated Lewis (RT1(1)) splenocytes. The Buffalo recipients were given 1 ml of rabbit anti-rat lymphocyte serum intra-peritoneally at the time of the donor antigen injection and 21 days later underwent a heterotopic 15-cm Lewis small bowel transplant. Pretransplant intrathymic Lewis alloantigen and anti-rat lymphocyte serum treatment prolonged Lewis small bowel survival by approximately 2.5 times (mean survival time = 18.0 vs 7.0 days for controls, P < 0.05). These small bowel allografts demonstrated pericryptic T cell infiltrates, areas of cryptic necrosis, and a dense submucosal lymphocytic infiltrate, all consistent with acute resection. In contrast to our studies achieving donor-specific cardiac allograft survival, this protocol did not result in indefinite intestinal allograft survival. No Buffalo recipients developed evidence of graft-versus-host disease.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Diagnosis and management of biliary complications of laparoscopic cholecystectomy.

Laparoscopic cholecystectomy has become the operation of choice for symptomatic cholelithiasis. However, this operation may result in serious biliary complications. Our aims were to review our experience with biliary complications of laparoscopic cholecystectomy and to document the mechanisms of the injuries and the techniques of managing these complications. We treated 20 patients with biliary complications of laparoscopic cholecystectomy. Symptomatic collections of bile (bilomas) were present in five patients. One of these patients underwent operative ligation of an accessory bile duct in the gallbladder bed, whereas the others had percutaneous or endoscopic therapy. In the remaining 15 patients (of whom 13 were referred from other hospitals), injuries to the major bile ducts were managed by combined radiologic, endoscopic, and operative therapies. In 10 of these patients (67%), the mechanism of injury was the misidentification of the common bile duct as the cystic duct. In 3 of 15 patients, a noncircumferential injury to the lateral aspect of the common bile duct occurred. The Bismuth levels of the remaining bile duct injuries were type I in 3, type II in 4, type III in 3, and type IV in 2. Early outcome of therapy for these bile duct injuries has been favorable. One patient was lost to follow-up, and 2 died of nonbiliary causes, whereas 12 patients are alive and well with normal serum liver enzyme levels at 4 to 19 months postoperatively (mean: 14 months). The most common cause of major bile duct injury during laparoscopic cholecystectomy is mistaking the common bile duct for the cystic duct. Most bilomas can be managed successfully with noninvasive methods. Coordinated efforts by radiologists, endoscopists, and surgeons are necessary to optimize the management of patients with major bile duct injury, suggesting that patients with biliary complications of laparoscopic cholecystectomy should be referred to specialty centers for optimal care.

Abdominal Pain↗
PubMed1993

Advantages of donor-specific blood transfusion in the rat via the portal vein for renal transplantation.

It is well known that blood transfusion can promote immune unresponsiveness to a renal allograft in both animals and humans. The present study examined the differences between either portal venous (PV) or intravenous (IV) donor-specific transfusion (DST) alone on the survival of rat renal allografts without other immunosuppression. Seven days before LEW (RT1(1)) renal transplantation, recipient WKA rats (RT1u) were inoculated with graded doses (1.0 to 0.0001 mL) of LEW whole blood through either the portal vein or the tail vein. A dose of 0.001 mL whole blood PV resulted in significantly greater prolongation of renal allograft survival than occurred with IV preimmunization (38.5 +/- 10.6 days v 12.5 +/- 2.7 days). Although recipient survival was not significantly different for the two routes for a lower dose (0.1 mL) of whole blood preimmunization 7 days before transplantation, azotemia and rejection were prevented when the PV route, but not the IV route, was used. However, when 0.1 mL of whole blood was administered 5 days before transplantation, survival was prolonged only in animals preimmunized by the PV route (54.7 +/- 14.0 days v 14.3 +/- 4.0 days). In the stronger reciprocal combination of WKA to LEW, there was an advantage of PV with 0.1 mL whole blood, but not with 1 mL. These results indicate that the beneficial effects of DST on renal allograft survival can be obtained when a smaller volume of blood is given via the PV rather than the IV route and a shorter interval elapses between preimmunization and transplantation.

Animals↗
PubMed1993

Preservation of renal function in juxtarenal and suprarenal abdominal aortic aneurysm repair.

PURPOSE: Deterioration in renal function is a common cause of morbidity in patients treated surgically for juxtarenal and suprarenal abdominal aortic aneurysms. We reviewed our experience over the last 8 years with 65 consecutive patients undergoing juxtarenal (n = 31) or suprarenal (n = 34) abdominal aortic aneurysm repair. METHODS: The aneurysms were repaired with a transabdominal (n = 8), thoracoabdominal (n = 4), retroperitoneal (n = 22), or thoracoretroperitoneal (n = 31) approach. Proximal aortic clamps were placed at the suprarenal, supra-superior mesenteric artery, or supraceliac level. Renal hypothermia with cold heparinized saline solution renal artery perfusion was used to protect renal function in 38 patients with either preoperative renal insufficiency or with anticipated prolonged renal ischemia (> 30 minutes). Concomitant renal artery reconstruction was required in 30 patients. RESULTS: Significant operative morbidity developed in 23 (35.3%) patients. There was one (1.53%) perioperative death (0 to 90 days). Temporary dialysis was necessary in two patients. Preoperative renal insufficiency was a significant risk factor on multivariate analysis for a decline in renal function during the first postoperative week. However, serum creatinine concentration had returned to baseline or improved in all patients but two (3.1%) at the time of discharge. In spite of significantly longer renal ischemia, discharge creatinine levels were, on univariate analysis, statistically less than baseline creatinine levels in patients with suprarenal aneurysms, patients requiring renal reconstruction, and patients treated with renal hypothermia. The location of the proximal aortic clamp was not a factor in postoperative morbidity. There was no significant difference between juxtarenal and suprarenal aneurysms with respect to operating room time, transfusion requirements, days intubated, resumption of oral diet, or the length of hospitalization. CONCLUSIONS: Careful consideration of the route of exposure, location of the proximal aortic clamp, and the preservation of renal function with renal hypothermia and with the repair of significant renal artery lesions will result in minimal morbidity and mortality in patients requiring surgery for juxtarenal or suprarenal abdominal aortic aneurysms.

Aged↗