The combination chemotherapy including cis-diamminedichloroplatinum (CDDP), adriamycin (ADM) and 5-fluorouracil (5-FU) is reported as one of the most effective regimens of urothelial cancer. We experienced one patient who had multiple metastatic urothelial cancer in the lung, liver and brain and who showed complete response, even in short duration, by this regimen. Since then, we have been using this regimen as the adjuvant chemotherapy of the invasive urothelial cancer. This regimen consists of 15 mg/m2 CDDP on days 1 to 5, 30 mg/m2 ADM on day 1 and 300 mg/m2 5-FU on days 1 to 5, and is repeated 3 times for 3 to 4 weeks. If toxicity is intolerable, the dosage of CDDP and ADM was decreased in thirty percent. As the maintenance, tegaful was administrated oral or suppository for 1 to 2 years. Seven cases which were followed for at least 12 months were evaluated. In one patient, local recurrence appeared after 16 months, but the other patients were disease free. General malaise, nausea, vomiting and alopecia were recognized in all patients in various degrees, but severe myelosuppression did not appear. Because of the toxicity, three patients had to have reduced dosage. In the adjuvant chemotherapy the most effective regimen should be used in the first therapy to eradicate the micrometastasis. At present, the combination chemotherapy of CDDP and ADM is the most effective for urothelial cancer as reported by many authors. Although the number of patients and the duration of follow up are inadequate to evaluate this adjuvant chemotherapy, we believe that this regimen is effective since invasive urothelial cancer usually recurs within 2 years.

Aged↗

PubMed1985

Translocation of proteins into rat liver mitochondria. Existence of two different precursor polypeptides of liver fumarase and import of the precursor into mitochondria.

Two different putative precursor polypeptides of rat liver fumarase were synthesized when RNA prepared from rat liver were translated in vitro using the rabbit reticulocyte lysate system. One of these putative precursor polypeptides (P1) was synthesized as a larger molecular mass than the mature subunit of fumarase (45,000 daltons) by about 5,000 daltons and the other (P2) had the same molecular mass as the mature enzyme. When the 35S-labeled cell-free translation products were incubated with rat liver mitochondria at 30 degrees C, P1 and the 35S-labeled mature size fumarase were associated with the mitochondria. Of these, the 35S-labeled mature size fumarase was resistant to externally added protease, but P1 was not, indicating that the 35S-labeled mature size fumarase was located in the mitochondrial matrix. The following observations strongly suggested that the 35S-labeled mature size fumarase in mitochondria was derived from P1, which was energy-dependently imported and concomitantly processed to the mature size. 1) The amount of the 35S-labeled mature size fumarase recovered from the mitochondria increased proportionally to the duration of incubation, while the amount of P1 recovered from the post-mitochondrial and mitochondrial fractions decreased with the duration of the incubation. 2) Only P1 could bind with the mitochondrial outer membrane at 0 degrees C even in the presence of an uncoupler of the oxidative phosphorylation but P2 did not. 3) P1 bound to the mitochondrial outer membrane was imported into the matrix, when the mitochondria binding only P1 at 0 degrees C was reisolated and incubated at 30 degrees C in the presence of an energy-generating system. The specific receptor was involved in the binding of P1 to mitochondria, since a high concentration of NaCl did not interfere with the binding of P1 to the membrane and did not discharge P1 bound onto the membrane. It was shown that P1 formed an aggregate composed of 6 to 8 molecules and P2 was a dimer in the cell-free translation mixture and that P1 and P2 were enzymatically inactive. These results suggest that the precursor for the mitochondrial enzyme has a larger molecular weight than that of the mature enzyme, whereas the precursor for the cytosolic enzyme has the same molecular weight as the mature enzyme.

Animals↗

PubMed1985

A rapid isolation technique of unmodified human T cells on a polystyrene resin column.

An easy and rapid isolation technique of human T cells on a polystyrene resin particle column has been developed. The cells of the effluent fraction contained more than 90% sheep erythrocyte (SRBC) rosette-forming cells and less than 1% of cells bearing surface immunoglobulin (Ig) or peroxidase. The T cell (SRBC rosette-forming cells) recovery rate was 80%. The distribution of OKT antigen T cell subsets was essentially the same as that of T cells separated by rosette sedimentation. Cell functions such as tritiated thymidine uptake by T cells and helper activity in Ig production were also the same as that of T cells separated by SRBC rosette sedimentation. Natural killer-like activity of the T cells isolated by the present method increased more than that of T cells obtained by the conventional method. Moreover, it was free from functional modification which tends to result from stimulation such as by the SRBC antigen in the SRBC centrifugation method. The combination of a T cell population offered by the present method and B cells depleted of SRBC-binding B cells minimized background plaque formation and enabled us to quantify the plaque-forming cell number in an antigen-specific plaque-forming assay. Furthermore, these populations produced relatively pure interleukin 2 (IL 2) by stimulation of an autologous mixed lymphocyte reaction without any absorption of IL 2 produced in the same culture. It seemed to be useful to evaluate the ability of lymphocytes from normal individuals and patients to produce IL 2.

Antigens, Surface↗

PubMed1985

Nature of the suppressor cells mediating prolonged graft survival after administration of extracted histocompatibility antigen and cyclosporine.

Antigen-specific suppressor T cells are induced by donor histocompatibility antigen extracted from spleen cells with 3M KCl combined with cyclosporine (Ag-CsA). A single i.v. injection of 5 mg 3M-KCl-extracted donor Buffalo (Buf, RT1b) antigen (Ag) combined with a three day course of CsA prolonged renal allograft survival in Wistar-Furth (WFu, RT1u) hosts to a greater extent (MST 26.5 days) than CsA alone (MST 11.8 days). Peripheral blood lymphocytes (PBL) or spleen cells harvested from Ag-CsA-treated recipients ten days after transplantation inhibited the mixed lymphocyte reaction (MLR) between normal responder WFu cells and irradiated Buf cells (55.6% and 64.4% suppression, respectively, P less than 0.025), but not third-party Brown-Norway (BN, RT1n) stimulator cells (13.6% and -18.3% suppression, respectively, NS). The suppressor effect was not mediated by cytolytic cells; there was neither primary nor secondary cytolytic activity against 51Cr-labeled Con-A blastoid Buf cells. The suppressor cells were neither adherent to plastic dishes nor to nylon-wool columns. PBL irradiated with 800 rads, but not 1500 rads, suppressed the MLR. A single injection of cyclophosphamide (CY, 25 mg/kg) seven days after transplantation abrogated the suppression induced by Ag-CsA treatment. Moreover, PBL from Ag-CsA recipients failed to suppress the MLR, if depleted either of all T cells by treatment with monoclonal antibody (Mab) W3/13 HLK (pan T cells; % suppression -15.8), or of cytotoxic/suppressor cells with Mab OX-8 (-19.3% suppression) together with rabbit antimouse immunoglobulin and complement. On the other hand, PBL treated with the Mab W3/25 (helper) showed suppressor cell activity (+56.4%, P less than 0.001) similar to untreated cells (62.4%, P less than 0.001). Moreover, adoptive transfer of suppressor T cells purified from pooled lymphocytes by rosetting using Mab significantly prolonged the survival of donor-specific, but not third-party, test grafts in naive secondary hosts. Thus, these studies demonstrated antigen-specific suppressor T cells mediate the long-term unresponsiveness induced by the Ag-CsA regimen.

Animals↗

PubMed1985

The immunosuppressive action of suppressor cells from antigen-cyclosporine-treated hosts on renal allograft survival.

Systemic adoptive transfer was employed to assess the immunosuppressive efficacy of antigen-specific suppressor T (Ts) cells purified from recipients treated with 3M KCl-extracted donor histocompatibility antigen (Ag) and cyclosporine (CsA). Suppressor cells were obtained from Wistar-Furth (WFu, RT-1u) hosts treated with a single i.v. injection of 5 mg 3M KCl-extracted donor Buffalo (Buf, RT-1b) antigen combined with a three-day course of CsA, a group that displays prolonged renal allograft survival (MST 23.2 +/- 10.2 days) compared with animals treated with CsA alone (MST 12.2 +/- 2.4 days). These noncytolytic, OX-8 phenotype, 800-rad-resistant/1500-rad-sensitive, nylon-wool-nonadherent and cyclophosphamide-sensitive suppressor T cells (1 X 10(6)) were adoptively transferred ten days after transplantation into virgin, secondary syngeneic hosts-thereby prolonging Buf graft survival from 7.2 to 17.5 days. The suppressor effect was immunologically specific; adoptive transfer did not prolong the survival of third-party Brown-Norway (BN) grafts (MST 10.4 +/- 3.1 days) compared with the nontreated control group (MST 11.0 +/- 2.9 days). The potency of Ts cells purified from Ag-CsA-treated hosts to transfer unresponsiveness into normal secondary WFu hosts (MST 17.5 +/- 8.0 days) was stronger than that of Ts cells from hosts treated with CsA only (MST 10.6 +/- 2.6 days). Moreover, in vitro stimulation of monoclonal-antibody-purified Ts cells by irradiated donor Buf spleen cells potentiated the in vivo induced suppressor activity, leading to an MST of 38.1 +/- 32.6 days; indeed 3 of 12 animals (25%) displayed permanent unresponsiveness. Furthermore, Ts cells from Ag-CsA-treated hosts displayed a synergistic effect with a three-day course of CsA administration into the secondary hosts (MST 24.2 +/- 8.0 days) compared with animals only treated with CsA (MST 12.2 +/- 2.4 days, P less than 0.001). Moreover, the combination of the Ag-CsA regimen with Ts cells administered one day after transplantation caused even greater prolongation of graft survival (MST 34.2 +/- 14.2 days) compared with Ag-CsA-treated hosts (MST 23.2 +/- 10.2 days, P less than 0.025). Thus adoptively transferred antigen-specific suppressor T cells may be explored to intensify the specific immunosuppressive effect of the Ag-CsA regimen to achieve long-term unresponsiveness.

Animals↗

PubMed1985

Pharmacodynamic assessment of the in vivo cyclosporine effect on interleukin-2 production by lymphocytes in kidney transplant recipients.

There are presently no monitoring tools to assess the immunosuppressive effect of cyclosporine (CsA) in vivo, since the mode of drug action is incompletely understood in man. Experimental in vitro studies suggest that CsA causes reversible inhibition of T helper cell generation of interleukin-2 (IL-2). Therefore the present study examined the effect of CsA administered in vivo on the capacity of kidney transplant recipient lymphocytes to generate IL-2 after mitogen (phytohemagglutinin [PHA]) stimulation. IL-2 production was measured by the capacity of lymphocyte supernates to trigger proliferation of a human IL-2-dependent T cell line. Peripheral blood lymphocytes (PBL) from CsA-Pred treated recipients displayed 40.6% inhibition (1.14 +/- 0.06 U/ml, n = 117, P less than 0.001) of IL-2 production compared with normal individuals (1.93 +/- 0.04 U/ml, n = 164). Dialysis patients did not display inhibited IL-2 production. The inhibition of IL-2 generation was observed in patients treated solely with CsA without supplemental corticosteroids (1.24 +/- 0.12 U/ml, n = 25; 35.8% inhibition, P less than 0.001). CsA did not inhibit the expression of the IL-2 receptor: 4.15 +/- 11.2% and 63.1 +/- 10.3 of normal lymphocytes and 36.7 +/- 9.8% and 60.4 +/- 12.2% of CsA-treated patient lymphocytes expressed anti-IL-2 receptors after 24 or 48 hr of PHA stimulation, respectively. Serial posttransplant studies in individual patients confirmed no inhibition of IL-2 generation pretransplant (1.94 +/- 0.07 U/ml) followed by a high degree of inhibition thereafter, namely 0.87 U/ml (55.0% inhibition) at 1 week, 1.15 U/ml (40.6% inhibition) at 2 weeks, 0.42 U/ml (78.2% inhibition) at 3 weeks, and 0.99 U/ml (48.7% inhibition) at 4 weeks. There was a correlation between the occurrence of rejection episodes in the 7 patients who suffered this event, and IL-2 generation by patient PBL. Before pulse therapy there was no inhibition of IL-2 generation (2.39 U/ml; -23.8 inhibition), documenting a poor level of immunosuppression in these patients. At 1, 2, 3, or 4 days after corticosteroid pulse therapy, PBL displayed 39.4%, 57.0%, 50.0% and 49.2% inhibition, respectively. These findings suggest not only that CsA treatment impairs the generation of IL-2 by patient lymphocytes, but also that failure to display this response is associated with a poor level of immunosuppression and allograft rejection. These studies provide a foundation for serial analyses of IL-2 generation, in order to dissect its utility as a pharmacodynamic parameter to assess the level of CsA-induced immunosuppression.

Azathioprine↗

PubMed1985

Suppressor cell activity of cells infiltrating rat renal allografts prolonged by perioperative administration of extracted histocompatibility antigen and cyclosporine.

Suppressor T cells were identified in situ within renal allografts of hosts rendered unresponsive by perioperative administration of donor histocompatibility antigen, which was extracted from donor spleen cells with 3M KCl, combined with cyclosporine (Ag-CsA). Infiltrating cells harvested from Buffalo (BUF, RT1b) renal allografts ten days after transplantation into Wistar-Furth (WFu, RT1u) rats treated with a single i.v. injection of 5 mg 3M KCl-extracted donor antigen (Ag) combined with a three day course of CsA inhibited the mixed lymphocyte culture (MLC) reaction between normal responder WFu and irradiated BUF cells (79.3% suppression, P less than 0.001), but not third-party Brown-Norway (BN, RT1n) stimulator cells (-5.4% suppression, NS). The suppressor effect was not due to cytolysis: the infiltrating cells did not lyse 51Cr-labeled concanavalin A (Con-A) blastoid BUF cells, as did the infiltrating cells from nonimmunosuppressed recipient allografts undergoing rejection responses toward BUF (49% specific cytolysis, E/T = 25), but not third-party BN, target cells five days after transplantation. The suppressor cells were nonadherent to plastic dishes and sensitive to monoclonal antibodies (Mab) W3/13 HLK (pan-T cells: % suppressor -17.9) or cytotoxic/suppressor cells with Mab OX-8 (-5.0% suppression), but not W3/25 (helper; 48.6% suppression, P less than 0.025). Moreover, adoptive transfer of 10(6) infiltrating cells from Ag-CsA-treated recipient allografts into virgin WFu hosts prolonged primary BUF graft survival from 7.2 to 14.0 days (P less than 0.05), but not third-party BN grafts (treated MST = 11.9 +/- 3.9 days versus untreated MST = 11.0 +/- 2.9 days, NS). On the other hand, infiltrating cells from CsA-only-treated recipient allografts could not transfer this effect (MST = 7.7 +/- 0.5 days, P less than 0.01). Finally, retransplantation of the BUF allograft from the Ag-CsA treated rat to a syngeneic, virgin WFu host ten days after primary transplantation yielded prolonged survival with MST 11.4 +/- 2.3 versus control primary graft survival in untreated animals of 7.2 +/- 0.6 days (P less than 0.001). BUF allografts from treated WFu hosts retransplanted into third-party BN rats did not display prolonged graft survival (MST = 9.2 +/- 1.1 days) compared with primary BUF grafts in untreated BN recipients (MST = 9.2 +/- 2.0 days, NS). The presence of suppressor T cells both in the spleen and in situ in renal allografts following Ag-CsA treatment suggests that local mechanisms may augment systemic elements to control the generation of alloimmunity.

Animals↗

PubMed1985

Metabolism of fentanyl in isolated hepatocytes from rat and guinea pig.

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Animals↗

PubMed1985

[Effect of hyperthermic irrigation with bleomycin on prophylaxis of recurrence of bladder cancer].

Hyperthermic irrigation with Bleomycin of bladder at 43 approximately 44 degrees C for 3 hours after operation was performed in about 80 cases of bladder cancer from 1978, in our hospital. Bleomycin (90 mg) in 3,000 ml saline solution, was irrigated for 10 days postoperatively. No severe side effect was observed, while the complaint of a local side effect (bladder irritability) was made in 57.8%. Non-recurrence rate was calculated by the actual method, in 54 patients with bladder cancer which occurred initially. Non recurrence rate was 88.0% after 1 year, 73.3% after 2 and 3 years, and 63.5% after 4 years or more. Recurrence rate per 100 patient-month was 0.98. On the basis of our observations, it was concluded that hyperthermic irrigation with Bleomycin was useful for the prophylaxis for recurrence of bladder cancer after removal of the tumor.

Adult↗

PubMed1985

Intracellular localization of low and high calcium-requiring forms of calpain*.

Monospecific and discriminative antibodies against two distinct forms of calpain (Ca2+-dependent cysteine proteinase) were obtained. Immunological cross-reactivity between the heavy subunits of calpain I (low Ca2+-requiring form) and calpain II (high Ca2+-requiring form) were calculated to be 15 to 17%, and two steps of affinity chromatography was required to obtain discriminative antibodies. Using these antibodies, we have shown that a minor but reproducible amount of calpain I is associated with the membrane of human erthrocytes. Indirect immunofluorescent staining of cultured PK 15 cell revealed diffuse and finely granular cytoplasmic staining. Porcine kidney stained with either anti-calpain I IgG or anti-calpain II IgG yielded strong stainings in proximal and distal convoluted tubules, while glomerulus, macula densa and interstitial cells were not stained.

Animals↗

PubMed1985

[A case of hypothyroid myeloneuropathy with giant axon--on the findings of sural nerve biopsy].

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Humans↗

PubMed1985

[Calpain and calpastatin in pig retina].

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Animals↗