Individually distinct tumor-specific cell surface antigen identified by monoclonal antibody on a Rous sarcoma virus-induced mouse tumor.

Hybrid cell lines were prepared by the fusion of BALB/c myeloma P3U-1 cells with the lymphocytes of BALB/c mice that were immunized with syngeneic Rous sarcoma virus (RSV)-induced tumor CSA1M cells. Three clones of the hybrid progeny (3.4B2, 3.4C6, and 3.5C11) produced cytotoxic IgM antibodies against CSA1M cells. One of the clones, 3.5C11, was chosen for analysis of the detailed specificity. Both direct cytotoxicity assays and absorption tests revealed that monoclonal antibody from 3.5C11 was positive only with CSA1M cells and that it failed to react with other tumors, including 20 RSV-induced mouse tumors, and normal cells. The 3.5C11 monoclonal antibody alone, with or without exogenous complement, was suppressive in the therapy of ip injected CSA1M tumor in syngeneic hosts, and significant prolongation in survival was seen in the treated mice. These results clearly showed presence of an individually distinct tumor-specific cell surface antigen on an RSV-induced mouse tumor.

Partial purification of a growth factor synthesized by a rat hepatoma cell line established in serum-free medium.

A rat hepatoma cell line was established from primary culture using RPMI 1640 without supplements. Hepatomas were induced in rats by 0.06% 3'-methyl-4-dimethylaminoazobenzene. An established cell line, FF101, has been maintained as a monolayer for longer than 34 months and subcultured for 42 passages. The population-doubling time was 78 h. The modal chromosome number was 66. FF101 was transplantable, and morphological examination of the transplanted tumors revealed a mixed type of hepatocellular and cholangiocellular carcinoma. FF101 retained the ability to express tyrosine aminotransferase and glucose-6-phosphatase. Also, FF101 synthesized alpha-fetoprotein. FF101-conditioned medium stimulated DNA synthesis and proliferation of several cell lines such as AH66, K562, and BALB/c3T3. The growth-promoting activity of FF101-conditioned medium was abolished by protease, dithiothreitol, acidic treatment, and heating. Gel filtration of conditioned medium on Sephacryl S-200 disclosed the growth-promoting activity at the molecular size of approximately 60,000 Da, and the isoelectric point (pI) was between 5.5 and 6.5. These results suggest that FF101 synthesizes a novel growth factor which has little specificity in both species and organs.

Transcriptional regulation of Mesp1 and Mesp2 genes: differential usage of enhancers during development.

Mesp1 and Mesp2 encode bHLH-type transcription factors, Mesp1 and Mesp2, respectively. The expression of both genes is observed in the nascent mesoderm, and subsequently in the rostral presomitic mesoderm. To determine the regulatory mechanism for gene expression, we attempted to identify enhancer elements by transient transgenic analysis. At least two enhancers, which are responsible for the expression of the two genes in the early mesoderm (early mesodermal enhancer, EME) and the presomitic mesoderm (PSM enhancer, PSME), and one suppressor, which is responsible for the rostrally restricted expression in the presomitic mesoderm, were identified. Deletion studies of these enhancer elements indicate that either gene may use the same enhancer for early mesoderm development, whereas both genes may utilize separate enhancers to regulate their expression in the presomitic mesoderm.

A synthetic peptide homologous to retroviral envelope protein down-regulates TNF-alpha and IFN-gamma mRNA expression.

We investigated the influence of CKS-17, a synthetic heptadecapeptide that corresponds to a highly conserved domain of the immunosuppressive retroviral envelope protein p15E, on staphylococcal enterotoxin B (SEB)-induced TNF-alpha gene expression in human peripheral blood mononuclear cells and highly purified human monocyte preparations, as well as the production of TNF-alpha protein, using human peripheral blood mononuclear cells. RNA hybridization studies show that CKS-17 inhibits SEB-induced TNF-alpha mRNA accumulation in human peripheral blood mononuclear cells and human monocytes. CKS-17 is also shown to be highly suppressive for SEB-induced production of TNF-alpha proteins. Similarly, CKS-17 inhibits expression of SEB-induced IFN-gamma mRNA in human peripheral blood mononuclear cells. These results suggest that CKS-17 down-regulates both TNF-alpha and IFN-gamma production at mRNA level.

Immunosuppressive retroviral peptides: immunopathological implications for immunosuppressive influences of retroviral infections.

Studies of the effects of retroviruses on the immune system, which date back through thirty years of investigations, are reviewed. In the earliest published studies in the 1960s, it was demonstrated that mice infected with oncogenic viruses were immunosuppressed. Since then, numerous articles have been published describing profound immunodeficiencies observed in vivo in humans infected with human immunodeficiency virus and in animals such as cats infected with the feline immunodeficiency virus. In vitro investigations have shown that inactivated retroviruses or transmembrane envelope protein p15E as well as a synthetic 17-amino acid peptide (CKS-17) impressively conserved within the transmembrane envelope protein of several animal or human retroviruses are highly immunosuppressive. More recently, dysfunction of cytokines produced by CKS-17 at both a cellular and molecular level have been found to mimic influences observed in vivo in patients infected with the human immunodeficiency virus. CKS-17 has also been shown to induce cAMP in vitro. The significance of these observations to understanding the immunological disturbances observed in malignancy, cytokine biosynthesis, and modulations of immune functions through cAMP is discussed.

Enrichment of murine bone marrow natural suppressor activity in the fraction of hematopoietic progenitors with interleukin 3 receptor-associated antigen.

Natural suppressor (NS) activity has been identified in several sites of active hematopoiesis. In this study we characterized NS activity in murine bone marrow (BM) using monoclonal antibodies (mAbs) to interleukin 3 (IL-3) receptor-associated antigen (IL-3RAA) and various cytokines that exert a strong influence on hematopoiesis or lymphocyte interaction. NS activity of BM cells of relatively low density was enhanced by IL-3 or granulocyte-macrophage colony-stimulating factor (GM-CSF). When the BM cells were separated into IL-3RAA+ cells and IL-3RAA- cells, the IL-3RAA+ cells demonstrated potent NS activity, whereas IL-3RAA- cells had either no or weak NS activity. The IL-3RAA+ cells showed non-T- and non-B-cell phenotype and had high affinity to wheat germ agglutinin (WGA), a marker for hematopoietic progenitors. In assays for hematopoietic activity, it appeared that the early differentiating progenitors (day 8 spleen colony-forming units [CFU-S], granulocyte-macrophage colony-forming units [CFU-GM]) were enriched in the IL-3RAA+ cell population, whereas more immature multipotent progenitors (day 12 CFU-S, granulocyte erythrocyte macrophage megakaryocyte colony-forming units [CFU-GEMM]) were contained in the IL-3RAA- cell population. Both suppressor cells and IL-3RAA+ cells spontaneously developed from the IL-3RAA- cell population. These findings suggest that NS cells in murine BM are early hematopoietic progenitors and are probably committed to the myeloid lineage. Hybridoma cells established between the IL-3RAA+ cells and BW5147 cells produced suppressor factor(s). This finding suggests that the NS cells produce soluble mediator(s) that may be responsible for their suppressive action.

HIV-1 recombinant gp41 induces IL-10 expression and production in peripheral blood monocytes but not in T-lymphocytes.

The effects of recombinant gp41 (rgp41) protein of the human immunodeficiency virus type 1 (HIV-1) on interleukin 10 (IL-10) expression and production using human peripheral blood mononuclear cells was investigated. Expression of IL-10 mRNA was demonstrated within 3 h of cell exposure to endotoxin-free rgp41 by RT-PCR and Northern blot analyses in a time- and dose-dependent manner. IL-10 protein was detected in the supernatants of peripheral blood mononuclear cells following stimulation with rgp41 also in a dose dependent manner. Fractionation of peripheral blood mononuclear cells showed that purified monocytes but not purified T-lymphocytes induced expression of IL-10 mRNA by rgp41. Recombinant HIV-1 gp120 exhibits similar influences on the induction of IL-10. These results indicate that both of these components of envelope proteins may play an important role in HIV related immunomodulation by influencing regulatory functions of monocytes and macrophages.

Involvement of a herbimycin A-sensitive protein tyrosine kinase in extracellular action of HIV-1 Nef.

Extracellular Nef which has been implicated in disease progression and development of AIDS induces IL-10, a potent immunosuppressive cytokine, in vitro. The present study was designed to examine whether the action of extracellular Nef is a protein tyrosine kinase (PTK)-dependent event. Anti-phosphotyrosine immunoblotting reveals that recombinant HIV-1 Nef induces rapid tyrosyl phosphorylation of several cellular proteins in human peripheral blood mononuclear cells. Pre-treatment of cells with herbimycin A, but not with genistein, significantly abolishes the Nef-induced tyrosine phosphorylation of cellular proteins. Furthermore, ELISA and RNase protection assays show that herbimycin A significantly blocks Nef-induced production of IL-10 at both the protein and the mRNA level. Genistein and aminogenistein have a much less blocking effect on the ability of Nef to induce IL-10. These results provide evidence for the involvement of a herbimycin A-sensitive PTK in the signal transduction pathway for exogenous HIV-1 Nef.

Prolactin acts on the extreme 5' portion of MMTV LTR involving a mammary cell-specific enhancer.

We have previously shown that a human mammotropic polypeptide hormone, prolactin (PRL) can act synergistically with steroid hormones to regulate gene expression directed by the long terminal repeat of mouse mammary tumor virus (MMTV LTR) in a human ductal carcinoma cell line T47D cells using a chloramphenicol acetyltransferase reporter gene system and gene transfection methods. In the present study, using various recombinant plasmids we analyzed functional elements in the MMTV LTR that is essential for the PRL responses. We show that the PRL-responsive elements are located in the extreme 5' end of the MMTV LTR, a region previously described by others to be a mammary cell-specific enhancer.

Prolactin, epidermal growth factor or transforming growth factor-alpha activate a mammary cell-specific enhancer in mouse mammary tumor virus-long terminal repeat.

Mammary specific expression of elevated levels of mouse mammary tumor virus (MMTV) contributes to mammary carcinogenesis. Mechanisms which regulate provirus expression have not been completely defined. Using a MMTV-long repeat terminal (MMTV-LRT) directed chloramphenicol-acetyltransferase (CAT) reporter gene system and a human breast cancer cell line T47D, we demonstrate that prolactin (PRL), epidermal growth factor (EGF), or transforming growth factor-alpha (TGF-alpha) act on a mammary cell-specific enhancer at the extreme 5' end of the MMTV-LTR involving sequences -1094 through -858. PRL and either EGF or TGF-alpha exert concerted roles in this activation of these sequences. In contrast, using a plasmid construct lacking this mammary cell-specific enhancer, EGF or TGF-alpha, but not PRL, act synergistically with progesterone to induce CAT activity, indicating that the action of PRL on regulatory elements of the MMTV-LTR is restricted to this mammary cell-specific enhancer involving sequences -1094 through -858. A mobility shift assay was used to demonstrate that PRL, EGF or TGF-alpha induce nuclear factors (MP4, MAF, and MGF) which bind directly to this mammary cell-specific enhancer element.

Expression of alpha-smooth muscle actin in small bronchioloalveolar adenocarcinoma of the lung: assessment and comparison with noguchi criteria.

The Noguchi criteria are useful in assessing the prognosis of patients with small lung adenocarcinoma. Although there is a significant difference in prognosis between type A or B and type C, it is difficult in some cases to distinguish these types accurately by microscopy. In this study, we used immunohistochemistry to examine alpha-smooth muscle actin (alpha-SMA) produced by active fibroblasts in 25 pulmonary adenocarcinomas less than 2 cm in diameter. Eleven of type C (61%) showed positive staining for alpha-SMA, whereas no positive cases were seen in type A or B. The incidence of cancerous blood vessel and lymphatic invasion were significantly higher in alpha-SMA positive cases than in negative cases, and the positive cases showed poorer prognosis. These findings indicate that immunohistochemical detection of alpha-SMA is useful and essential for histological typing by the Noguchi criteria.

Repair of salivary fistula after reconstruction of pharyngoesophagus.

We describe three surgical methods used to repair salivary fistulas in different situations that are seen after reconstruction of the pharyngoesophagus in patients with hypopharyngeal cancer. The principle of these methods is the same; a cervical skin flap next to the fistula is used as the internal lining and an anterior chest skin flap is rotated as an external lining to cover it. For the primary closure of heavily irradiated fistulas, this combination may not always be effective, and we emphasize that a combined use of a pedicled pectoralis major muscle flap with these two kinds of pedicled skin flap is extremely reliable.

Effect of hyperthermia on the laryngeal closure reflex. Implications in the sudden infant death syndrome.

A recent study with dogs has defined an age-related period of transient hyperexcitability in the laryngeal adductor neurons. This observation suggests that age-related neurologic instability of laryngeal mechanisms and abnormal laryngeal closure may be a cause of transient upper airway obstruction, inducing potentially fatal central apnea that could result in the sudden infant death syndrome. Since hyperthermia has been implicated as a cofactor in sudden and unexpected infant death, its effect on laryngeal excitability requires clarification. This investigation has found that both the latency and threshold of the laryngeal adductor reflex decreases during hyperthermia. Thus, hyperthermia effectively enhances this reflex during its age-related period of hyperexcitability, and is capable of triggering upper airway-induced central apnea of sufficient severity to result in death. The effect of temperature on latency is attributed to changes in axonal conduction and synaptic transmission velocities. Temperature-dependent changes in synaptic transmission are hypothesized as the cause of the observed threshold changes.

Renovascular hypertension: a case of intimal fibroplasia with congenital anomaly of the renal artery.

We report a case of renovascular hypertension caused by fibromuscular dysplasia (intimal fibroplasia) with congenital anomaly of the renal artery. Arteriography revealed complete occlusion from the root to the middle of the right renal artery and a collateral artery from the right lumbar artery. Histological examination showed intimal fibroplasia and three arterioles in the intima of the renal artery.

Legionnaires' disease associated with habitual drinking of hot spring water.

A 57-year-old man presented with pneumonia, respiratory distress, and myelodysplastic syndrome. A diagnosis of Legionnaires' disease due to Legionella pneumophila (L. pneumophila) was established. The patient had long been drinking tap water via a conduit from a hot spring resource, from which L. pneumophila was also isolated. Both the patient's strain and the water strain of L. pneumophila were identified as serogroup 1, and the genetic relatedness between the two strains as seen by pulsed-field gel electrophoresis was 87%. The patient was successfully treated with erythromycin, fluoroquinolone, and rifampicin. This case raises an important issue on public health represented by legionellosis in Japan.

Comparison of changes in hemodynamics between unilateral and bilateral lung volume reduction for pulmonary emphysema.

This study was aimed to compare changes in hemodynamics between unilateral (UL) or simultaneous bilateral (BL) lung volume reduction surgery (LVRS) for chronic obstructive lung disease. Sixteen patients underwent LVRS by stapler resection with neodymium: yttrium-alminum-garnet (Nd: YAG) laser ablation; five underwent BL-LVRS (four by median sternotomy and one by thoracoscopy) and 11 underwent UL-LVRS by thoracoscopy. Four patients had multiple bullae within pulmonary emphysema. At preoperation and 6, 12, 24, and 48 hours postoperatively, hemodynamics and right ventricular performance were evaluated. UL- and BL-LVRS reduced afterload of the right and left ventricle postoperatively. Although the pulmonary arterial resistance increased after surgery, the total pulmonary resistance decreased (p=0.001) in association with the reduced systemic vascular resistance (p=0.001). These reductions improved cardiopulmonary circulation, resulting in increased stroke volume and cardiac output (p=0.003). The right ventricular ejection fraction showed minimal change 48 hours postoperation. Two patients died of pneumonia caused by persistent air leakage. In conclusion, both the UL- and BL-LVRS showed similar effectiveness in terms of improvement in the systemic and cardiopulmonary circulation after LVRS, if there were no postoperative complications. We concluded that we had to reduce and repair the persistent air leakage after LVRS.

Effectiveness of prostaglandin E1 on pulmonary hypertension and right cardiac function induced by single-lung ventilation and hypoventilation.

It is known that prostaglandin E1 (PGE1) is a potent vasodilator and improves red cell deformability. Single lung-ventilation sometimes occurs under lung transplantation, lung cancer surgery and traumatic pneumonectomy, and may result in increased pulmonary resistance, right heart failure and severe hypoxemia. The present experimental study was undertaken to examine the effects of PGE1 on these states induced by single-lung ventilation and hypoventilation. Fourteen pigs weighing 32-33 kg were anesthetized, intubated and ventilated using a respirator and then randomly assigned to two groups, the control group and the PGE1 treated group, 7 pigs each. After median sternotomy to induce severe hypoxemia hypoventilation was induced and then the right hilus pulmonis was cross clamped. Mean blood pressure, mean pulmonary arterial pressure (PAP), pulmonary vessel resistance (PVR), right ventricular stroke work (RVSW) and arterial blood gases (PaO2 and SatO2) were measured at baseline, in the hypoventilation state, and 15 min, 1 hour, and 2 hours after the right hilus pulmonis clamping with hypoventilation. PGE1 (250 microg/20 ml saline) was administered via the central vein starting 15 min after right hilus cross clamping for 1 hour and 45 min in the PGE1 group. PGE1 significantly reduced PAP and PVR, normalized RVSW, and improved PaO2. PGE1 may be useful for the condition of increased pulmonary hypertension during single-lung ventilation and hypoventilation.

Fibrinolytic activity in medium from tissue culture of paranasal mucous membrane.

It is known that a remarkable fibrinolytic activity of plasminogen activator (PA) can be seen in extracts of wet tissue and acetone powder preparation of paranasal mucous membrane evidencing chronic sinusitis. However, the origin of the PA in extract of paranasal mucous membrane has not yet been clarified up to the present time. In this experiment, using a tissue culture of paranasal mucous membrane, it was observed that two species of cells, epithelial cells and fibrocytes, proliferated in the implanted tissue. PA was isolated from the conditioned medium on the fifth day after culture. From these results, it appears that the PA may be released from epithelial cells and/or fibroblasts.

Some chemical properties of tissue plasminogen activator purified from paranasal mucous membrane.

Plasminogen activator (PA) was purified from an acetone powder preparation of paranasal mucous membrane with chronic sinusitis, and some chemical properties of the purified PA were investigated in this paper. Zn-imminodiacetate affinity chromatography, lysine sepharose affinity chromatography and ultrafiltration for concentrating a PA fraction were consecutively performed to purify the PA from the acetone powder preparation. Finally, gel filtration was performed using Sephacryl S-200 in order to estimate the molecular weight of the purified PA. The purified PA in this experiment showed a stronger affinity to fibrin than urokinase did. The molecular weight of the purified PA was estimated to be 65,000 to 70,000 daltons as determined by Sephacryl S-200 gel filtration. The Km of the purified PA was 0.11 mM. From these results, it is apparent that the PA purified from an acetone powder preparation of paranasal mucous membrane belongs to the class of tissue type plasminogen activators (t-PA).

Characteristics of the epithelial component of parotid adenolymphoma.

Parotid adenolymphoma is composed of two histologic components, epithelial and lymphoid. Although some theories regarding the histogenesis of this tumor have long been disputed, there have been no definite conclusions. The purpose of this study was to clarify the origin of the epithelial components of this tumor using histochemical and immunopathological techniques, electron microscopy and a survey of HE-stained tumor sections. The results obtained indicated that the functions of the epithelial components were similar to those of the striated duct of the normal parotid gland, and morphological studies showed that the origin of the epithelial components may arise from parotid ductal inclusion in the lymphnodes in or around the parotid gland.