Changes in the immunoreactivity for inhibin in mouse retina during development of photoreceptor cells

INTRODUCTION: A costovertebral paraganglioma with a long history of recurrences and metastases is reported. MATERIAL AND METHODS: The lesion is documented on histological ground and with immunocytochemistry and electron microscopy. RESULTS: The patient is a 42 year old man which presented with a 5 cm axillary mass and enlarged mediastinal lymph nodes. The former lesion has been interpreted as a metastasis of paraganglioma. The patient underwent surgery because of a costovertebral paraganglioma at the age of 19 and showed locoregional metastases and a recurrence over a period of 23 years. Twenty-seven years from removal of the primitive tumor, the patient is alive and well. All the neoplasms showed identical histological features as they were composed of neoplastic elements arranged in nests surrounded by dendritic cells. Neoplastic cells immunoreacted with anti-chromogranin antiserum whereas sustentacolar cells were positive with S-100 antiserum. DISCUSSION: The differential diagnosis between a multicentric paraganglioma a metastasis has been particularly taken into consideration. In addition, axillary tumor was distinguished from a metastatic renal cell carcinoma and melanoma. The literature regarding costovertebral paraganglioma has been also revised.     

CD28-independent, TRAF2-dependent costimulation of resting T cells by 4-1BB ligand

4-1BB ligand (4-1BBL) is a member of the tumor necrosis factor (TNF) family expressed on activated antigen-presenting cells. Its receptor, 4-1BB, is a member of the TNF receptor family expressed on activated CD4 and CD8 T cells. We have produced a soluble form of 4-1BBL using the baculovirus expression system. When coimmobilized on plastic with anti-CD3, soluble 4-1BBL induces interleukin (IL)-2 production by resting CD28+ or CD28- T cells, indicating that 4-1BBL can function independently of other cell surface molecules, including CD28, in costimulation of resting T cell activation. At low concentrations of anti-CD3, 4-1BBL is inferior to anti-CD28 in T cell activation. However, when 4-1BB ligand is provided together with strong TCR signals, then 4-1BBL and anti-CD28 are equally potent in stimulation of IL-2 production by resting T cells. We find that TNF receptor-associated factor (TRAF)1 or TRAF2 associate with a glutathione S-transferase-4-1BB cytoplasmic domain fusion protein in vitro. In T cells, we find that association of TRAF1 and TRAF2 with 4-1BB requires 4-1BB cross-linking. In support of a functional role for TRAF2 in 4-1BB signaling, we find that resting T cells isolated from TRAF2-deficient mice or from mice expressing a dominant negative form of TRAF2 fail to augment IL-2 production in response to soluble 4-1BBL. Thus 4-1BB, via the TRAF2 molecule, can provide CD28-independent costimulatory signals to resting T cells.     

Disulfide bond mapping and structural characterization of spruce budworm antifreeze protein

We have previously reported that bryostation 1 (Bryo 1) induces differentiation of chronic lymphocytic leukemia (CLL) in vitro to a hairy cell (HC) stage. This study tests the hypothesis that Bryo 1-differentiated CLL cells are more susceptible to 2-chlorodeoxyadenosine (2-CdA) than parent CLL cells. A recently established EBV-negative CLL line (WSU-CLL) from a patient resistant to chemotherapy including fludarabine was used to test this hypothesis. Both Bryo 1 (10-1000 nM) and 2-CdA (5.6-22.4 microM) exhibited a dose-dependent growth inhibitory effect on the WSU-CLL cell line. In vitro, the sequential exposure to Bryo 1 (100 nM for 72 h) followed by 2-CdA (11.2 microM) resulted in significantly higher rates of growth inhibition than either agent alone. Changes in immunophenotype, enzymes, lipids, proteins, and the DNA of WSU-CLL cells were studied before and after Bryo 1 treatment. Bryo 1 induced a positive tartrate-resistant acid phosphatase reaction and two important markers, CD11c and CD25, after 72 h of culture, confirming the differentiation of CLL to HC. The Fourier transformation infrared spectroscopic analysis showed that the amount of membrane lipids significantly increased in Bryo 1-treated cells compared to controls after 24 h, whereas the protein content, as well as the DNA content, decreased. This finding supports the change of CLL to HC. To evaluate the in vivo efficacy of Bryo 1 and 2-CdA, we used a xenograft model of CLL in WSU-CLL-bearing mice with severe combined immune deficiency. s.c. tumors were developed by injection of 10(7) WSU-CLL cells, and fragments were then transplanted into a new batch of severe combined immunodeficient mice. Bryo 1 and 2-CdA at the maximum tolerated doses (75 micrograms/kg i.p. and 30 mg/kg s.c., respectively) were administered to the mice at different combinations and schedules. The survival in days, the tumor growth inhibition ratio, the tumor growth delay, and the log10 kill of the mice treated with Bryo 1 followed by 2-CdA were significantly better than the control and other groups. We conclude that the sequential treatment with Bryo 1 followed by 2-CdA resulted in higher antitumor activity and improved animal survival.     

Cytotoxic potential of monoalkylation products between mitomycins and DNA: studies of decarbamoyl mitomycin C in wild-type and repair-deficient cell lines

Hypoxic regions in solid neoplasms have been associated with tumor recurrence and resistance to several cancer treatment modalities including radiation therapy. Various strategies have been designed to target these resistant cells, including the use of the bioreductive alkylating agent mitomycin C (MC), which exerts preferential cytotoxicity under hypoxic conditions in most cell lines. Analyses of the mechanism of action of MC indicate that this drug can form cross-links with DNA; it is currently thought that this bisadduct is the critical lesion responsible for inhibiting DNA synthesis. Computer-generated models suggest that the MC adduct fits snugly into the minor groove of B-DNA without imposing major distortion on the structure of the DNA molecule. To gain additional insight into the role of cross-linkage in the cytotoxicity of MC, we studied the analogue, decarbamoyl mitomycin C (DMC). The structure of DMC is identical to that of MC with the exception of the substitution of the carbamoyl group at the C-10 position by a nonalkylating hydroxyl group (-OH); this alteration would be expected to prevent DMC from forming bisadducts with DNA. In chemical systems, DMC produces only DNA monoadducts. If indeed it is the MC-DNA cross-links which are responsible for cell kill, one would predict DMC to be less cytotoxic than MC. However, tissue culture studies using DMC revealed that DMC is at least as toxic as MC to EMT6 mouse mammary tumor cells and to wild-type AA8 Chinese hamster ovary (CHO) cell lines.(ABSTRACT TRUNCATED AT 250 WORDS)     

Identifying sites of replication initiation in yeast chromosomes: looking for origins in all the right places

DNA fragments that contain an active origin of replication generate bubble-shaped replication intermediates with diverging forks. We describe two methods that use two-dimensional (2-D) agarose gel electrophoresis along with DNA sequence information to identify replication origins in natural and artificial Saccharomyces cerevisiae chromosomes. The first method uses 2-D gels of overlapping DNA fragments to locate an active chromosomal replication origin within a region known to confer autonomous replication on a plasmid. A variant form of 2-D gels can be used to determine the direction of fork movement, and the second method uses this technique to find restriction fragments that are replicated by diverging forks, indicating that a bidirectional replication origin is located between the two fragments. Either of these two methods can be applied to the analysis of any genomic region for which there is DNA sequence information or an adequate restriction map.     

Local anesthetic block of batrachotoxin-resistant muscle Na+ channels

Local anesthetics (LAs) are noncompetitive antagonists of batrachotoxin (BTX) in voltage-gated Na+ channels. The putative LA receptor has been delineated within the transmembrane segment S6 in domain IV of voltage-gated Na+ channels, whereas the putative BTX receptor is within segment S6 in domain I. In this study, we created BTX-resistant muscle Na+ channels at segment I-S6 (micro1-N434K, micro1-L437K) to test whether these residues modulate LA binding. These mutant channels were expressed in transiently transfected human embryonic kidney 293T cells, and their sensitivity to lidocaine, QX-314, etidocaine, and benzocaine was assayed under whole-cell, voltage-clamp conditions. Our results show that LA binding in BTX-resistant micro1 Na+ channels was reduced significantly. At -100 mV holding potential, the reduction in LA affinity was maximal for QX-314 (by 17-fold) and much less for neutral benzocaine (by 2-fold). Furthermore, this reduction was residue specific; substitution of positively charged lysine with negatively charged aspartic acid (micro1-N434D) restored or even enhanced the LA affinity. We conclude that micro1-N434K and micro1-L437K residues located near the middle of the I-S6 segment of Na+ channels can reduce the LA binding affinity without BTX. Thus, this reduction of the LA affinity by point mutations at the BTX binding site is not caused by gating changes induced by BTX alone. We surmise that the BTX receptor and the LA receptor within segments I-S6 and IV-S6, respectively, may align near or within the Na+ permeation pathway.     

IFN-tau: a novel subtype I IFN1. Structural characteristics, non-ubiquitous expression, structure-function relationships, a pregnancy hormonal embryonic signal and cross-species therapeutic potentialities

IFN-tau (IFN-tau) constitutes a new class of type I IFN which is not virus-inducible, unlike IFN-alpha and IFN-beta, but is constitutively produced by the trophectoderm of the ruminant conceptus during a very short period in early pregnancy. It plays a pivotal role in the mechanisms of maternal recognition of pregnancy in ruminants and it displays high antiviral and antiproliferative activities across species with a prominent lack of cytotoxicity at high concentrations in vitro in cell culture and possibly in vivo. It exhibits high antiretroviral activity against HIV and exhibits immunosuppressive activity in a multiple sclerosis model and reduces embryo and fetal mortality by stimulation of IL-10 production. In this review all the biochemical and para-hormonal properties of this novel IFN-tau are described in detail: structural characteristics of proteins and genes, trophoblast expression, regulation of its expression, structure of its gene promoter, its absence in human species and in non-ruminant animals, the evolution of the IFN-tau genes, its structure-function relationships with its three-dimensional structure, structural localization of biological activities, its lack of cytotoxicity and its receptor. Surprisingly, for an IFN, IFN-tau is also a pregnancy-embryonic signal with paracrine antiluteolytic activity. In order to maintain luteal progesterone secretion, IFN-tau inhibits PGF-2alpha pulsatile secretion and oxytocin uterine receptivity in early pregnancy. It is believed to suppress pulsatile release of endometrial PGF-2alpha by preventing oxytocin and estrogen receptor expression. Additionally, it directly regulates prostaglandin metabolism and possibly the PGE:PGF-2alpha ratio.