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Biomedical subjects

F Vari

Publications and source records attributed to F Vari.

At least 19 recordsLinked to original sources
PubMed2004

Loading DCs with Ag.

Understanding the biology of the DC and its pivotal role in immune response initiation and regulation has enabled new effective anti-cancer therapies to be developed for treatment of a wide range of tumor types. Many studies on DC-based cancer vaccine development have focused on the development of methods that can effectively deliver tumor Ags to DCs. Numerous methods have been developed or evaluated for the delivery of defined and undefined tumor Ags to DCs for processing and presentation to T lymphocytes. This review provides a brief summary of these methods, the techniques that are used in each, how they have -- or could -- be applied clinically, as well as the advantages and disadvantages of each method.

Adoptive Transfer↗
PubMed1999

Immunization with a synthetic peptide conjugate derived from the N-terminal sequence of either the beta-chain of haemoglobin or the immunosuppressive protein (reOLT 4) reduces the litter size of pregnant rats.

A synthetic peptide conjugate based on the N-terminal sequence of a 10 000 MW immunosuppressive serum protein (reOLT 4) was used to immunize female Lewis rats prior to mating, in order to determine whether blocking this protein had an effect on pregnancy. The N-terminal sequence of (reOLT 4) has close sequence homology to the beta-chain of rat haemoglobin so a peptide conjugate based on the N-terminal sequence of this protein was also used to immunize female Lewis rats. Controls included animals that were not immunized and animals that received the peptide carrier, diphtheria toxoid (DT). No statistical differences were found in gestation time or litter sizes in these groups. Differences were, however, evident between these groups and animals that received DT-(reOLT 4) (group 4) or the DT-beta-chain haemoglobin (group 5). There were no statistical differences in litter size or gestation time for group 4 when compared with group 5. Enzyme-linked immunosorbent assay and dot-blot analysis revealed that rats from both groups also had strong responses against DT, the peptide conjugate they were immunized with and the corresponding full-length protein. In both cases, animals from group 4 and group 5 had weak responses to the peptide that they did not receive, together with lower erythrocyte counts and haematocrits, and elevated heart to body weight ratios. Additionally, antibody purified on a (reOLT 4) immunoaffinity column was capable of binding to rat erythrocytes. A second investigation comparing anaemia prior to fertilization and maintained anaemia over the gestation period revealed that only the latter was capable of decreasing litter size to the same degree as obtained for groups 4 and 5. We conclude that for groups 4 and 5 it is the autoimmune effect of continual anaemia over the gestation period, mediated by autoantibodies, which results in the observed lower litter size.

Animals↗
PubMed1998

Analysis of immunosuppressive proteins in serum of liver-transplanted rats by using an anti-LSF-1 affinity column.

Liver suppressor factor one (LSF-1) is a 40-kDa immunosuppressive protein in the serum of rats 60 days after orthotopic liver transplantation (OLT) between the nonrejector combination of DA donors into PVG recipients. In the present study, the purification of proteins from rat OLT serum taken 60 days after transplantation was performed by affinity chromatography using the anti-LSF-1 polyclonal antibody (pAb). The assessment of column eluates using anti-LSF-1 and OLT serum was studied using rat heart and liver transplantation models. Rejection was not suppressed by the administration of OLT serum in heart or liver allografts. However, heart allografts treated with peak eluates (450 micrograms single shot im, dissolved in Intralipos) taken from the affinity OLT serum survived significantly longer than untreated rats (median = 36.5 days; n = 7 vs 6.5 days; n = 5, respectively, P = 0.011). The same treatment with anti-LSF-1 column eluates also prolonged liver allografts significantly (>200 days) than those in either the untreated group (median = 11 days; n = 7) or those which received only Intralipos (median = 10.5 days; n = 5, P = 0.019). Subsequent analysis of the N-terminal sequences of some of the proteins which were eluted from the affinity column revealed that the homology of a 30-kDa protein was identical to hemoglobin alpha-chain, a 59-kDa protein to granulocyte inhibitory factor, a 70-kDa and a 90-kDa to albumin and its precursor, respectively. Although the specific immunosuppressive component has not been isolated, our results suggested that the anti-LSF-1 column can extract immunosuppressive moiety of LSF-1 from OLT serum.

Animals↗
PubMed1997

The characterization of reconstituted passenger leukocytes on the induction of tolerance in rat liver transplantation.

The tolerance induced by orthotopic liver transplantation [DA (RT1a) rats to PVG (RT1c) rats] can be prevented by total body irradiation of the donor rat. Reconstitution of the irradiated donor with DA splenic leukocytes reintroduces this tolerance. To investigate the major histocompatibility complex (MHC) specificity of passenger leukocytes, irradiated DA donors were reconstituted by third-party BN (RT1n) splenic leukocytes. The reconstitution with BN splenocytes re-established DA-specific tolerance in PVG recipients, as confirmed by subsequent DA cardiac allografting, while BN hearts were rejected with second-set tempo. To determine which cell components play an important role in re-establishing liver graft tolerance, DA splenic leukocytes were further purified into three types: T, B, and adherent cells. Only "T-cell-enriched" preparations restored liver graft tolerance in three out of five PVG recipients. These results suggest that passenger leukocytes of differing MHC types can help to induce liver-specific tolerance and that T cells in the liver graft may be essential to regulate tolerance induction.

Animals↗
PubMed1997

The prevention of graft-versus-host disease by the serum of liver retransplanted rats.

The effect of serum from orthotopic liver retransplanted rats (re-OLT serum) on graft-versus-host disease (GVHD) was studied in rats. In the re-transplantation model of rat liver, orthotopic liver transplantation (OLT) was carried out in the DA (RT1a) into PVG (RT1c) combination; two days later the DA liver was removed and a new PVG liver implanted into the same recipient (re-OLT). In the in vivo GVHD model, male PVG rats were sublethally irradiated and injected intravenously with 3 x 10(8) DA or BN (RT1n) spleen through the penial vein. Within 1 h of the inoculation, rats of the experimental group were injected with 1 ml of re-OLT serum taken at postoperative day (POD) 7. Rats in the control group received 1 ml of normal PVG serum or syngeneic re-OLT serum (PVG-PVG, PVG-PVG). All PVG rats in the control groups died of GVHD within 21 days after the inoculation of DA or BN spleen lymphocytes. However, when the animals were treated with re-OLT serum, 100% (6/6) of the rats survived more than 60 days, following inoculation with DA lymphocytes but not with BN lymphocytes. The POD 7 re-OLT serum showed a strong inhibition against DA anti-PVG mixed lymphocyte reaction (MLR), although re-OLT serum did not contain soluble DA class I antigens, anti-DA class I or II antibody. The potential GVHD inhibitory factors in re-OLT serum may be two unique immunosuppressive proteins, which have been detected by SDS PAGE and reported previously. We conclude that re-OLT serum has immunosuppressive factors, which, at least in part, prevented the induction of GVHD in rats.

Animals↗
PubMed1997

Differences in the rate of donor leucocyte migration between natural and drug-assisted tolerance following rat liver transplantation.

The number involved in and the rate of migration of donor leucocytes into the following recipient organs (spleen, thymus, bone marrow, lung and mesenteric lymph nodes) were measured in two rat models of orthotopic liver transplantation (OLT) using donor-specific MHC class I antibodies. The first OLT model is one that does not require immunosuppression in order to achieve tolerance and involved the transplantation of DA (MHC haplotype, RT1a) livers into PVG (RT1c) recipients. The second model was one that required a 7-day (10 mg/kg) treatment with cyclosporin A (CsA) to achieve tolerance and used DA donors into Lewis (RT1(1)) recipients. Recipient organs were biopsied on days 3, 20 and 87 following OLT and donor leucocyte migration was quantified by immunohistochemistry and computer densitometry of immunoblots of detergent-solublized tissues in order to resolve both membrane-bound and soluble donor MHC class I antigen. In a separate experiment, spleen biopsies were taken following OLT on days 3 and 15 from the naturally tolerizing OLT model (DA into PVG), treated with and without CsA for 7 days and compared with the (DA into Lewis) model. The initial rate of leucocyte migration between days 3 and 21 following OLT was found to be the most rapid into the spleen, followed by the bone marrow and mesenteric lymph nodes in the naturally tolerant (DA into PVG) model when compared with the (DA into Lewis) model. The number of donor leucocytes in the spleen and mesenteric lymph nodes in both models was, however, approximately the same by 87 days. No real difference in the rate of leucocyte migration was seen in the thymus or the lung for both transplant models over the time course assayed. CsA was found to lower the rate of donor leucocyte migration only over the period it was administered. The rate of donor leucocyte migration into the spleen was still much lower 15 days after OLT in the (DA into Lewis) model compared with the (DA into PVG) model treated with and without CsA. Thus the differences in the rate of donor leucocyte migration into the spleen, bone marrow and mesenteric lymph nodes immediately following OLT may offer an explanation as to why the (DA into PVG) combination is able to accept a transplanted liver without immunosuppressive therapy.

Animals↗
PubMed1996

Novel immunosuppressive proteins purified from the serum of liver-retransplanted rats.

Liver grafts between certain rat strain combinations, such as DA (RT1a)-into-PVG (RT1c), are accepted without the use of immunosuppressive agents. To explore the nature and role of serum proteins in liver-induced immunosuppression, we have developed a retransplantation model of rat liver grafting. In this procedure, orthotopic liver transplantation (OLT) is carried out in the DA-into-PVG combination; two days later the DA liver is removed and a new PVG liver implanted into the same recipient (re-OLT). Serum from re-OLT rats was immunosuppressive when tested in mixed lymphocyte reactions (MLR). Three novel proteins were detected in re-OLT serum by SDS-PAGE, with sizes of 180 kD, 87 kD, and 10 kD. The N-terminal sequences of these were distinct and did not match protein sequences in the computer databases, although there was some homology between the 10 kD sequence and the beta-chain of rat hemoglobin. Purified 87 kD and 10 kD proteins were immunosuppressive in MLR; in both cases suppression was dose-dependent and nonstimulator-specific. Production of the 180 kD and 87 kD molecules required the presence of the recipient spleen. We conclude that re-OLT serum contains novel immunosuppressive proteins, which may be products of immune recognition and associated with the immediate termination of graft rejection.

Amino Acid Sequence↗
PubMed1996

A simplified silver diammine method for the staining of nucleic acids in polyacrylamide gels.

High sensitivity and low background, which are the significant features of the procedure of Johansson and Skoog (J. Biochem. Biophys. Methods. 1987, 14, (Suppl.) 33) for silver staining of nucleic acids in polyacrylamide gels, have been improved by excluding ethanol from all solutions and increasing the length of washing steps after sensitisation and staining. Including a fixation step before sensitisation increased the contrast of the bands even at high concentration of nucleic acid, producing a shift in the tone of colour from black to grey instead of gold to yellow bands. This technique is superior in terms of sensitivity and background to a number of other silver staining protocols, three which utilise silver diammine and three which utilise silver nitrate ions.

Ammonia↗
PubMed1996

Restoration of tolerance to rat hepatic allografts by spleen-derived passenger leukocytes.

The tolerance induced by orthotopic liver transplantation (OLT) in certain combinations of rat strains can be prevented by total body irradiation (TBI) of the donor. We demonstrate here that the intravenous inoculation of splenic leukocytes into irradiated donors before OLT could re-establish tolerance in association with a state of microchimerism detected in the recipients. When donor DA (RT1a) strain rats were irradiated with 1000 rad 24 h before liver harvesting and subsequent liver implantation into PVG recipients, five out of six rats died from rejection in this normally tolerogenic OLT (DA-PVG) combination. Injection of 1.5 x 10(8) splenic leukocytes from naive DA rats into the irradiated DA donor rats 24 h before OLT restored the tolerogenic potential of the liver allografts. Immunofluorescence assay revealed an increased number of donor (DA) type cells in the PVG recipient bearing a repopulated DA liver, compared to the PVG recipient of an irradiated liver. These results suggest that passenger leukocytes reconstituted by splenic leukocytes have the capacity to protect liver allografts.

Animals↗
PubMed1996

The beneficial effect of the prostacyclin analogue (OP 2507) on rat liver transplantation subjected to an extended anhepatic phase.

We investigated the effect of a prostacyclin analogue (OP 2507) on PVG (RT1c) recipients subjected to an extended anhepatic phase (AH) and transplanted orthotopically with PVG livers. All of the animals that underwent orthotopic liver transplantation (OLT) with a 20-min AH survived for 1 week with or without OP 2507, (OP) treatment (10/10, 100%). When the AH was lengthened to 45 min, the 1-week survival rate of recipients was poor in the OP-untreated groups (1/10, 10%). Treatment of the recipient with OP 2507, 0.15 micrograms/kg per minute for 30 min, prior to the 45-min AH substantially improved 1-week survival (5/6, 83.3%, P < 0.05). The serum TNF-alpha level at day 1 in OP-treated animals that underwent OLT with a 45-min AH was significantly lower than that in animals with 45-min AH OLT without OP treatment. We conclude that OP 2507 treatment has potential usefulness as a perioperative treatment when the AH is extended during OLT.

Animals↗
PubMed1996

Allograft acceptance and rejection, mediated by a liver suppressor factor, LSF-1, purified from serum of liver transplanted rats.

In certain rat strain combinations liver allografts are spontaneously accepted without immunosuppression and induce donor-specific tolerance to further skin and heart graft in the recipient. Such an effect is also transferrable using serum from orthotopically liver transplanted rats (OLT serum). In the OLT serum of one such combination, DA (RT1a) donor into PVG (RT1c) recipient, a 40 kDa protein (liver suppressor factor, LSF-1) has been identified and shown to be immunosuppressive in vitro. The aim of the present study is to investigate the immunological effect of LSF-1 and a polyclonal antibody (anti-LSF-1) against this molecule, in a rat heterotopic heart transplant (HHT) model and OLT model, respectively. Intramuscular injection of 300 micrograms of LSF-1, 1 h postoperatively, into a PVG recipient of either a DA or BN (RT1n) cardiac allograft caused significant prolongation of graft survival. Intravenous injection of polyclonal rabbit sera raised against an N-terminal peptide of LSF-1 (anti-LSF-1), within 1 h postoperatively, had variable effects on the survival of DA liver grafts in PVG recipients. In 5/6 cases injection of between 1 and 2 ml of anti-LSF-1 resulted in death of the recipient. Histological examination of the liver showed severe rejection with lymphoid cell infiltration of the portal tract and sinusoids and extensive damage to the parenchyma. All control rats survived for more than 60 days without any signs of rejection. The anti-LSF-1 polyclonal antibody prevented the injection of tolerance in the normally tolerogenic model (DA into PVG). This, together with the in vivo results, suggests a role for LSF-1 in the induction of tolerance.

Animals↗
PubMed1995

Orthotopic liver retransplantation in rats.

A surgical experience with a method of rate orthotopic liver retransplantation (OLRT), and a preliminary study of immunological responses after OLRT are reported. OLRT was performed on the same recipient after the fist orthotopic liver transplantation (1st-OLT) according to our original (Kamada's) cuff method. Replacement of the portal vein (PV) and infra-hepatic vena cava (IHVC) cuffs was not technically difficult. However, there were no survivors from the first 6 retransplanted rats, mainly due to complications from defective supra-hepatic vena cava (SHVC) anastomoses. Unlike the human intra-abdominal SHVC, the posterior wall of the intra-abdominal SHVC in rats is too short and fragile to perform an end-to-end anastomosis twice between donor and recipient SHVC. For a second group of seven retransplants, a modification of the SHVC anastomosis was made between donor and recipient SHVC in conjunction with the recipient's cuff diaphragm. This enabled reanastomosis to be secure, resulting in the improved 1-week survival after isogenic OLRT (85.7%). This OLRT model has been applied to the fully allogeneic combination for several immunological studies and led to novel findings. Thus, an experimental model of a rat orthotopic liver retransplant model has the potential to allow more valuable insights into the immunological study of chronic rejection, sensitization and chimerism following liver retransplantation.

Anastomosis, Surgical↗
PubMed1987

Defects in tumor cell immunogenicity: lymphokine signals in in vitro cytolytic responses to tumor alloantigens.

The B16 melanoma of C57BL/6 mice illustrates a deficiency in immunostimulation which may be important in some host-tumor relationships. B16 immunizes very poorly, even against its own major histocompatibility complex (MHC) antigens. We have compared the anti-MHC cytolytic response induced in vitro by B16 and by other tumors of both lymphoid and nonlymphoid origin. We have also studied the role of indomethacin and exogenous lymphokines in facilitating these responses and examined the relationship of specific and nonspecific effector cells induced. In contrast to normal lymphoid cells and two lymphoid tumor cells (EL4 and WEHI-265), the three nonlymphoid tumors, B16, Lewis lung tumor (3LL), and MC-2 fibrosarcoma, failed to induce primary cytolytic responses by themselves. MC-2 and B16 represented two different defects in immunogenicity. MC-2, which we have shown previously to induce an in vivo cytolytic response, could also immunize in vitro provided that prostaglandin production was blocked with indomethacin. In contrast B16, which is poorly immunogenic in vivo, immunized in vitro only if a concanavalin A-induced lymphokine supernatant (CS) was added as an exogenous source of "signal 2." High concentrations of the interleukin 2-containing Con A-induced spleen cell culture supernatant-induced non-H-2b-specific lymphokine-activated killer (LAK) cells in the absence of B16 stimulator cells. However, lymphokine concentrations too low to induce LAK cells enabled the otherwise nonimmunogenic B16 cells to induce specific cytolytic activity.

Animals↗