Damage accrual related to pregnancies before and after diagnosis of systemic lupus erythematosus: a cross-sectional and nested case-control analysis from a lupus registry.

OBJECTIVE: The objective of this study was to evaluate the chronic damage associated with pregnancies before and after the diagnosis of systemic lupus erythematosus (SLE). METHODS: Using childbearing-aged female SLE patient data registered at the Okayama and Showa University Hospitals, a nested case-control analysis was performed to investigate the relationship between pregnancy and chronic damage using the Systemic Lupus International Collaborating Clinics/American College of Rheumatology Damage Index (SDI). RESULTS: Pregnancy occurred in 22 patients before and 13 patients after the diagnosis of SLE in 104 eligible patients. Live births occurred in 82% (33/40) and 50% (9/18) of the pregnancies before and after the diagnosis of SLE, respectively. After matching age and disease duration, 33 case patients with chronic damage (SDI ≥ 1) and 33 control patients without chronic damage (SDI = 0) were selected. Hypertension was more frequent in cases than in controls (48% vs. 24%, p = 0.041). Pregnancies before and after the diagnosis of SLE were comparable between cases and controls (before the diagnosis: nine case patients and eight control patients; after the diagnosis: three case patients and five control patients; p = 1.00). Even after adjusting for hypertension using multivariate analysis, the pregnancies before and after the diagnosis were not significant predictors for chronic damage (odds ratio = 1.48 (95% confidence interval 0.33-6.65)), p = 0.60 of the pregnancy before the diagnosis; odds ratio = 0.78 (95% confidence interval 0.13-4.74), p = 0.78 of the pregnancy after the diagnosis). CONCLUSION: Pregnancies, either before or after the diagnosis of SLE, did not show any differences in chronic damage. Our results help alleviate fears regarding childbearing in female patients with SLE and their families.

Progressive reduction of serum complement levels: a risk factor for relapse in patients with hypocomplementemia in systemic lupus erythematosus.

OBJECTIVE: Serologically active clinically quiescent (SACQ)-SLE is a subtype of systemic lupus erythematosus (SLE); most SACQ-SLE patients relapse. Although complement and/or anti-dsDNA level fluctuations during SACQ status are reportedly not useful for predicting relapse, they might be useful in specific clinical settings. We aimed to assess the correlation between future relapse and progressive reductions in serum complement levels following remission in patients with hypocomplementemia . METHODS: We retrospectively reviewed patients aged ≥15 years who were treated with ≥20 mg/day of prednisolone for remission induction. After achieving remission, the patients treated with prednisolone tapered to ≤15 mg/day without relapse and followed by hypocomplementemia (first hypocomplementemia point) were analyzed. The primary outcome was the relapse during the first 24 months. RESULTS: Seventy-six patients were enrolled; 31 (40.8%) relapsed. A ≥10% reduction after the first hypocomplementemia point in serum C3, C4, and CH50 levels was found in 10, 21, and 16 patients, respectively. Hazard ratios (95% confidence intervals) for relapse were 2.32 (0.92-5.12) for serum C3 levels and 2.46 (1.18-5.01) for serum C4 levels. Progressive reductions in serum C3 and C4 levels had relatively high specificity (93.3% and 82.2%) but limited sensitivity (22.6% and 41.9%) for predicting relapse. However, simultaneous progressive reduction in C3 levels and increase in anti-dsDNA antibody levels had the highest specificity (97.8%), and simultaneous progressive reduction in C4 levels or increase in anti-dsDNA antibody levels had the highest sensitivity (71.0%). CONCLUSION: Simultaneous progressive reductions in complement levels and increases in anti-dsDNA antibody levels may indicate future relapse SACQ-SLE patients.

Induction and properties of cytoplasmic factor(s) which enhance nuclear nonhistone protein phosphorylation in lymphocytes stimulated by anti-Ig.

An increased in vitro phosphorylation of nonhistone nuclear proteins (NHP) was observed in the nuclei isolated from rabbit lymphocytes which had been stimulated with anti-Ig for 4 h. No concomitant increase of phosphorylation in histones or 0.14 M NaCl-soluble proteins was observed. The increase of in vitro phosphorylation of NHP was also observed in the nuclei isolated from nonstimulated cells when these nuclei were preincubated for 2 h with cell-free extracts from anti-Ig-stimulated cells. The active substance in cell-free extracts was maximally induced when lymphocytes were stimulated with anti-Ig for 2 h. The induction of an increased phosphorylation of NHP in nonstimulated nuclei with the cell-free extracts was not due to decrease of the adenosine triphosphate pool in the extracts from anti-Ig-stimulated cells. The active substance in cell-free extracts was not NHP-protein kinase itself, but it probably activated NHP-protein kinase in quiescent nuclei. The active substance was nondialyzable and probably protein. It was resistant against heating at 56 degrees C for 30 min, but the activity was completely destroyed by heating at 90 degrees C for 30 min. The active substance may be responsible for the transduction of the membrane-mediated signals given through Ig receptors to nuclei.

B cell growth factors and B cell differentiation factor from human T hybridomas. Two distinct kinds of B cell growth factor and their synergism in B cell proliferation.

Human T hybridomas secreting B cell growth factors (BCGF) and B cell differentiation factor (BCDF) have been established. Hybrid clones 77-A, 94-C, and 98-F secreted BCGF that induced proliferation of anti-IgM-stimulated normal B cells. The culture supernatant from 77-A cells could also maintain continuous proliferation of colony-forming B cells, but the factor from 94-C could not. The addition of the supernatant from 94-C cells to that from 77-A cells, however, synergistically augmented the proliferation of colony-forming B cells, demonstrating the existence of two distinct kinds of BCGF and the synergism between them. These supernatants, however, showed no interleukin 2 (IL-2) or BCDF activity. A hybrid clone, 90-E, secreted BCDF. The culture supernatant induced Ig production in Cowan I-stimulated normal B cells or in a transformed B cell line, CESS. However, the supernatant had no BCGF or IL-2 activity. Anti-Ig-stimulated B cells, but not IL-2-dependent T cells, absorbed BCGF activity and CESS cells absorbed BCDF activity but not BCGF activity in the culture supernatants from T hybridomas. Taken collectively, the results demonstrated that IL-2, BCGF, and BCDF were different molecules and acceptors specific for the each molecule are present on the each target cell.

Hemoglobins M: identification of Iwate, Boston, and Saskatoon variants.

Hemoglobin M variants, M Iwate, M Boston, M Saskatoon are easily and accurately identified by electron spin resonance with small amounts of patients' blood. In hemoglobin M Iwate and M Boston the electron spin resonance of both fresh blood (unprocessed) and isolated pure ferrihemoglobin M revealed similar signal shapes; whereas that of hemoglobin M Saskatoon was doublet in fresh blood and triplet in pure ferrihemoglobin M.

OPC-21268, an orally effective, nonpeptide vasopressin V1 receptor antagonist.

An orally effective, nonpeptide, vasopressin V1 receptor antagonist, OPC-21268, has been identified. This compound selectively antagonized binding to the V1 subtype of the vasopressin receptor in a competitive manner. In vivo, the compound acted as a specific antagonist of arginine vasopressin (AVP)-induced vasoconstriction. After oral administration in conscious rats, the compound also antagonized pressor responses to AVP. OPC-21268 can be used to study the physiological role of AVP and may be therapeutically useful in the treatment of hypertension and congestive heart failure.

Potent aquaretic agent. A novel nonpeptide selective vasopressin 2 antagonist (OPC-31260) in men.

Solute-free water diuretics (aquaretics) by antagonizing hydrosmotic vasopressin receptors (V2) may be useful in treating water-retaining diseases. The effects of intravenous administration of a newly developed nonpeptide, selective V2 antagonist, OPC-31260, at doses ranging from 0.017 to 1.0 mg/kg to groups of healthy, normally hydrated men were compared with those of 0.33 mg/kg furosemide and placebo. OPC-31260 increased the hypotonic urine volume dose dependently for the first 4 h, while furosemide induced sodium diuresis for 2 h. The absolute increase in the cumulative response in the urine to the highest doses of OPC-31260 was not significantly different from that to furosemide. The higher doses of OPC-31260 rapidly lowered urine osmolality for 2 h, particularly between minutes 15 and 45 (e.g., 1.0-mg/kg dose: 63 +/- 2 mOsm/kg in urine collected between minutes 30 and 45). In a marked hypotonic diuresis, mean free water clearance of the 4-h urine increased dose proportionally into the positive range, reaching 1.80 +/- 0.21 ml/min at 1.0 mg/kg. Whereas furosemide induced marked Na and K diuresis, OPC-31260 increased urinary Na excretion only slightly. At 4 h, 0.75 and 1.0 mg/kg of OPC-31260 almost doubled the plasma arginine vasopressin; and the higher doses increased plasma osmolality and plasma Na slightly, but did not alter plasma K, blood pressure, or heart rate. OPC-31260 thus safely induced a potent aquaretic effect in men.

Increased frequency of antigen-specific CD8(+) cytotoxic T lymphocytes infiltrating an Epstein-Barr virus-associated gastric carcinoma.

Gastric adenocarcinomas carrying Epstein-Barr virus (EBV) are known to be accompanied by massive lymphocyte infiltration. To characterize the tumor-infiltrating lymphocytes (TILs), we isolated and cultured such cells from a surgically resected EBV-associated gastric carcinoma. They were found to be positive for CD3, CD8, T-cell receptor beta chain, and cytotoxic molecules. The isolated TILs consisted of human leukocyte antigen (HLA) class I-restricted CD8(+) cytotoxic T lymphocytes (CTLs), which killed autologous EBV-transformed cells (but not phytohemagglutinin blast cells) and recognized HLA-A24 as restriction molecules. However, the TILs did not recognize known EBV antigenic peptides presented by HLA-A24 molecules, nor HLA-A24(+) fibroblasts infected with vaccinia recombinant virus expressing each of the EBV latent proteins. EBV(+) gastric carcinomas do not express conventional target proteins of EBV-specific CTLs, and the data suggest that some cellular proteins may be involved in the strong T-cell response to EBV-associated gastric carcinoma. In addition, our data suggest that class I-restricted, antigen-specific CD8(+) CTLs are specifically expanded within EBV(+) gastric carcinoma tissue.

Synthesis of a novel cytokine and its gene (LD78) expressions in hematopoietic fresh tumor cells and cell lines.

The gene coding for the protein LD78 was isolated from stimulated human tonsillar lymphocytes by differential hybridization. The gene product consisted of 92 amino acids with characteristics of cytokines. LD78 gene transcripts were detected in eight of eight fresh samples of cells from patients with acute nonlymphocytic leukemia (ANLL) by Northern blot analysis. ANLL cells with monocytic features gave the strongest bands. RNA transcripts were found in two of three samples of cells from patients with adult T cell leukemia (ATL), eight of nine samples from patients with acute lymphocytic leukemia (ALL) of B cell lineage, and one of the three samples from patients with T cell ALL. KG-1, HL-60, HUT 102, MT-2, and MJ cell lines expressed the LD78 gene constitutively. The LD78 protein was detected in culture supernatants and cell lysates of HUT 102, MT-2, MJ, and fresh ATL cells by Western blot analysis. This protein was not found in culture supernatants or cell lysates of monocytic leukemia cells and HL-60 cells, although LD78 transcripts were found in those cells. The discrepancy between gene and protein expression might be explained by the stability of the mRNA. Thus, the protein may be involved in the neoplastic transformation of hematopoietic cells.

Erythropoietin and Friend virus gp55 activate different JAK/STAT pathways through the erythropoietin receptor in erythroid cells.

Abnormal erythropoietin (EPO)-independent cell growth is induced after infection of erythroid progenitor cells with a polycythemic strain of Friend virus (FVp). Binding of its Env-related glycoprotein (gp55) to the EPO receptor (EPOR) mimics the activation of the EPOR with EPO. We investigated the gp55-EPOR signaling in erythroblastoid cells from mice infected with FVp and in cells of FVp-induced or gp55-transgenic-mouse-derived erythroleukemia cell lines, comparing it with the EPO-EPOR signaling in EPO-responsive erythroblastoid cells. While the Janus protein tyrosine kinase JAK2 and the transcription factor STAT5 became tyrosine phosphorylated with the EPO stimulation in EPO-responsive erythroblastoid cells from anemic mice, JAK1 and STAT5 were constitutively tyrosine phosphorylated in all of these FVp gp55-induced erythroblastoid or erythroleukemic cells. Moreover, this constitutively tyrosine-phosphorylated STAT5 was unable to bind to its specific DNA sequences and did not translocate to the nucleus. Nuclear translocation and DNA binding of this STAT5 species required EPO stimulation. These findings clearly indicate that the FVp gp55-EPOR signaling is distinct from the EPO-EPOR signaling and suggest that STAT5 may not play an essential role in the transmission of the cell growth signals in FVp gp55-induced erythroleukemia cells.

Prevalence and correlates of cervical HPV infection in a clinic-based sample of HIV-positive Hispanic women.

OBJECTIVES: Puerto Rico (PR), is the fifth highest jurisdiction of the United States of America (US) with respect to HIV prevalence and the leading in cervical cancer incidence. This cross-sectional study describes the prevalence and correlates of cervical HPV infection among a clinic-based sample of 302 women living with HIV/AIDS in PR. METHODS: Data collection included questionnaires, blood and cervical samples. Multivariable logistic regression models were used to estimate the magnitude of association (adjusted Prevalence odds ratio [aPOR]) between HPV cervical infection and other covariates. RESULTS: Mean age of participants was 40.3 years (± 10.3SD). The prevalence of HPV infection was 50.3%; 41.1% for low-risk types and 29.5% for high-risk types. Having ≥ 10 lifetime sexual partners (aPOR = 2.10, 95% CI:1.02-4.29), an abnormal Pap (aPOR = 3.58, 95% CI:1.93-6.62), active genital warts (aPOR = 3.45, 95% CI:1.60-7.42), and CD4 counts ≤ 200 (aPOR = 4.24, 95% CI: 1.67-10.78) were positively associated with any cervical HPV infection. Similar results were observed for HR HPV infection. CONCLUSIONS: A high burden of HPV co-infection exists among women living with HIV/AIDS in this population. Given the high incidence of HIV in PR and the higher risk of cervical cancer among women living with HIV/AIDS, HPV vaccination should be promoted in this population.

Mediator production and tumorigenesis of murine monocyte lines.

A murine macrophage-like cell line P388D1 (PD1) and its subline PD2 were studied for mediator production and also for tumorigenic properties. When PD1 cells were cultured in a medium containing 5% fetal calf serum, the culture supernatants exhibited potent interleukin 1 (IL1) activity (also termed lymphocyte-activating factor). IL1 activity produced in unstimulated cultures was predominantly of a low-molecular-weight species (mol wt, 13,000-16,000), but in PD1 cultures stimulated with 10-20 microgram lipopolysaccharide/ml a major IL1 activity was associated with a substance(s) of 65,000-75,000 in molecular weight. The PD1 cell line was nontumorigenic in normal female, syngeneic, inbred DBA/2J mice but produced progressively growing tumors in animals that had been immunosuppressed by 450-rad whole-body X-irradiation and treatment with antithymocyte serum. Inoculation with PD1 failed to produce tumors in athymic nude mice, and the spleens of nude mice that rejected PD1 grafts contained significant levels of a theta-positive cell population(s). PD2, a nonadherent and nonphagocytic subline of PD1, produced very little IL1 but instead produced a potent lymphocyte-inhibiting factor in vitro. The PD2 cell line was highly tumorigenic in syngeneic DBA/2J hosts. These results indicated a direct association between tumorigenicity and the type of mediators produced by these murine macrophage cell lines.

Host responses with solid tumors. I. Monocytic effector cells within rat sarcomas.

Morphologic studies of two unrelated progressively growing sarcomas in rats revealed the presence of a large proportion of host cells distinct from tumor cells. Enzymatically or mechanically dispersed tumor suspensions were fractionated by the velocity sedimentation method, which resulted in a separation of host and tumor cells. The host cells had the typical morphology of macrophages and also were adherent to a plastic surface after a short incubation period. The proportion of host cells was greater in a young (10 or 12 days) than in an older tumor (35 days). These macrophage-like host cells isolated from sarcomas 12 and 35 days after implantation inhibited the colony formation of tumor cells in vitro in the nonspecific manner ascribed to activated marcrophages. In addition to these effector cells, the host fraction contained macrophage precursors able to proliferate in vitro but apparently inhibited by the presence of tumor cells. The results indicated that these macrophage-like effector cells may be the predominant host reaction within some rat sarcomas, and also that thymus- and bone marrow-derived lymphocytes act only indirectly if they are involved.

Dose, route, and time dependence of serum lysozyme and antitumor activity following administration of glucan, Corynebacterium parvum, pyran, or lipopolysaccharide to mice.

Following the administration of Di Luzio particulate glucan, Corynebacterium parvum, pyran (maleic anhydride vinyl ether 6), and lipopolysaccharide (Shigella) to inbred C57BL/6J mice, dose, route, and time-dependent studies were undertaken on antitumor activity and serum lysozyme levels to explore the possible relevance of serum lysozyme as a useful index of antitumor activity. Antitumor activity was assessed by measurement of the extent of loss of iv injected 125I-labeled 5-iodo-2'-deoxyuridine-labeled B16 tumor cells. Increases in serum lysozyme levels were dose-, route-, and time-dependent and varied greatly from one agent to another. The peak levels of antitumor activity were similar for all agents but were also critically dose- and time-dependent. Correlations of serum lysozyme levels and antitumor activity were inexact. The doses for peak lysozyme level increases were higher than those for peak antitumor activity. Antitumor activity peaked earlier and lasted longer than serum lysozyme level increases.

Inhibition of pulmonary metastasis by Nocardia rubra cell wall skeleton, with special reference to macrophage activation.

The antimetastatic activity of Nocardia rubra cell wall skeleton (N-CWS) with or without cyclophosphamide was examined in an experimental model of pulmonary metastasis induced by Lewis lung carcinoma in C57BL/6 mice. Lewis lung carcinoma cells were implanted into the footpads of mice, and the implanted tumors were removed 9 to 10 days later. Pulmonary metastatic nodules began to develop a few days after the implanted tumor was removed. The inhibitory effect of N-CWS was evaluated from the number of pulmonary surface nodules about 3 weeks after tumor implantation. The antimetastatic activity of N-CWS depended upon the dose, time, and route of its injection. Injection of N-CWS i.v. after removal of the implanted tumor caused the greatest inhibition of development of pulmonary metastases. Therapy with N-CWS plus cyclophosphamide prolonged significantly the survival of mice with metastases. The cytotoxic activities of peritoneal macrophages and macrophages in the lung against Lewis lung carcinoma cells were enhanced in mice treated with N-CWS. Injection i.v. of peritoneal macrophages activated with N-CWS inhibited pulmonary metastases. The role of macrophages in inhibition of micrometastasis in the lung is discussed.

Association of macrophage activation with antitumor effect on rat syngeneic fibrosarcoma by Nocardia rubra cell wall skeleton.

The antitumor activities of the cell wall skeleton (CWS) of Nocardia rubra were demonstrated for syngeneic fibrosarcoma (AMC-60) in ACI/N rats in regard to macrophage activation. In the 24-hr cytolytic test, activated macrophages which were fractionated from peritoneal exudate cells induced by i.p. injection of Nocardia CWS showed significant cytolytic activity for [125I]iododeoxyuridine-labeled tumor cells. Activated macrophages also strongly inhibited [3H]thymidine incorporation into the tumor cells during the 24-hr cytostatic test. When tumor cells were inoculated s.c. with activated macrophages in the Winn-type transfer assay, subsequent tumor growth was significantly inhibited. Repeated i.p. injection of the CWS seemed to enhance these antitumor activities of macrophages. The therapeutic effect of Nocardia CWS was assessed with the ascites tumor and with the solid tumor inoculated i.m. into the hind leg. In the former treatment, repeated i.p. injections completely prevented the accumulation of ascites fluid and resulted in prolongation of the survival period. The peritoneal macrophages harvested from these survivors had a strong cytolytic activity for tumor cells in the cytolytic test. In the latter treatment, repeated intratumoral injections inhibited the growth of primary tumor and prevented metastasis. Furthermore, peritoneal resident macrophages from these tumor-bearing rats treated intratumorally with the CWS were found to be cytolytic for tumor cells in the cytolytic test.

Effect of Nocardia rubra cell wall skeleton on T-cell-mediated cytotoxicity in mice bearing syngeneic sarcoma.

Cell-mediated cytotoxicity against syngeneic MC104 fibrosarcoma cells was detected in C57BL/6N mice 7 days after tumor inoculation in the hind foot. This cytotoxicity was undetectable by Day 14 in the Winn test using spleen and draining popliteal lymph node (DPLN) cells. Similar results were obtained with the 51Cr release assay following in vitro activation of these lymphoid cells with mitomycin C-treated tumor cells. The antitumor cytotoxicity was shown to be mediated by T-cells. Spleen but not DPLN cells from 14-day tumor bearers enhanced tumor growth in the Winn test, suggesting the presence of immunosuppressor cells in the spleen. Two intralesional injections of 50 microgram of cell wall skeleton (CWS) of Nocardia rubra on Days 2 and 7 resulted in apparent tumor growth inhibition and prolongation of the survival period of tumor bearers. DPLN cells from tumor bearers treated with N. rubra CWS exhibited significant recovery in the cytotoxicity tested on Day 14, whereas the recovery in that of spleen cells was not apparent. The cytotoxicity augmented by N. rubra CWS was specific to MC104 tumor cells and was shown to be mediated by T-cells. These cytotoxic T-cells were shown to be able to localize not only in DPLN but also in the spleen and tumor in mice receiving the intralesional immunotherapy with N. rubra CWS. These results suggest that T-cell-mediated cytotoxicity against syngeneic tumor can be augmented by N. rubra CWS and might play an important role in the systemic development of its antitumor effect, although the effector cell increase in the spleen might be suppressed by splenic suppressor cells during tumor growth.

Characterization of new human pancreatic cancer cell lines which propagate in a protein-free chemically defined medium.

Many human cancer cell lines which have been maintained in fetal bovine serum (FBS)-supplemented medium produce and secrete many substances such as transferrin, alpha 1-antitrypsin, alpha 2-macroglobulin, alkaline phosphatase, gamma-glutamyltranspeptidase, creatine kinase, carcinoembryonic antigen, alpha-fetoprotein, carbohydrate antigen 19/9, and cytokines including colony-stimulating factors and transforming growth factor, and further they may produce small amounts of unknown substances. Usually, small amounts of substances have to be concentrated as highly as possible for detection, but FBS interferes with this procedure. A protein-free culture system is an ideal method for detecting small quantities of substances which originate from cancer cells without interference by FBS. However, we were concerned that protein-free culture may interrupt the production of the substances which have been produced in FBS-supplemented medium. In this study, we investigated the productibility of 46 kinds of well-known substances in ten newly established cell lines derived from human pancreatic cancer. These cell lines were propagated in a protein-free non-FBS-supplemented medium. Of the ten cases, one cell line alone that was derived from acinal cell carcinoma propagated as a semisuspension; on the other hand, nine cell lines that were derived from ductal cell carcinoma propagated as monolayers without piling up. This method prolongs the doubling time, which is not affected by the addition of FBS. The spent media of these cell lines were collected aseptically after the removal of cell debris and concentrated by ultrafiltration using a Pericon cassette followed by lyophilization. Using 46 kinds of available antibodies, we investigated whether or not the substances which react to these antibodies could be detected in the spent media and in the cells by enzyme-linked immunosorbent assay, Western blot analysis, and immunocytochemistry. Among these cell lines, HPC-Y11 produced and secreted the most kinds of substances, and the production of those substances was lowest in HPC-Y0. In conclusion, our protein-free culture system can be available in every laboratory, since this is not only an economical method, but also an effective method for the saving of purification procedures. Moreover, this is a most suitable method for surveying unknown substances derived from cancer cell lines.

Effect of Nocardia rubra cell wall skeleton on murine interferon production in vitro.

Nocardia rubra cell wall skeleton (N-CWS) induced alpha- plus beta-interferons (IFN-alpha/beta) in resident peritoneal cells and bone marrow cells from normal mice. N-CWS also induced IFN-alpha/beta and IFN-gamma in the cultures of thioglycollate-elicited peritoneal cells. When induced with N-CWS, the mixed cultures of thioglycollate-elicited macrophages and spleen cells produced high titered IFN-gamma. Both macrophages and lymphocytes present in spleen cells (non-T, non-B, asialo-GM1-positive) were essential for the production. Immunofluorescence staining with anti-IFN-gamma antiserum showed the presence of IFN-gamma in lymphocytes but not in macrophages, suggesting that lymphocytes were producer cells and macrophages were accessory cells. Because IFN-gamma is an important lymphokine in immune response, the ability of N-CWS to stimulate IFN-gamma production may account for a significant portion of the antitumor activity of this agent.

Induction of a tumor necrosis factor-like activity by Nocardia rubra cell wall skeleton.

Nocardia rubra cell wall skeleton (N-CWS) stimulated adherent cells harvested from the peritoneal cavities of thioglycollate-treated mice to produce cytotoxic activity. Depletion of macrophages from the adherent cells by 2-chloroadenosine or silica abrogated the production of this cytotoxic activity, whereas treatment of the adherent cells with anti-Thy-1.2 antibody and complement did not. This suggested that macrophages were the producer cells of the activity. Cytotoxic activity became detectable as early as 2 h after N-CWS treatment and reached peak activity at 9 h, then declined to a lower level, indicating rapid onset without persistent effects. N-CWS-induced cytotoxic factors have a fairly narrow temperature range, pH optimum for storage, and are sensitive to pronase and trypsin. By using column chromatography, N-CWS-induced cytotoxic factors were compared in detail with tumor necrosis serum obtained from Bacillus Calmette-Guérin endotoxin-treated mice. Both toxins were found to be nearly identical with respect to their behavior in ion-exchange, gel filtration, and concanavalin A affinity columns. N-CWS also induced human peripheral blood lymphocytes to release cytotoxic activity. Monocytes predominantly participated in production of this activity as confirmed by treatment with monoclonal antibody and complement. The cytotoxic activity was completely neutralized by anti-human tumor necrosis factor antiserum, but not by anti-human lymphotoxin antiserum. The fact that human peripheral blood lymphocytes release tumor necrosis-like factors after stimulation with N-CWS might account for the antitumor effects of this agent.