Fluorescence guided optical coherence tomography for the diagnosis of early bladder cancer in a rat model.

PURPOSE: We describe the technique of fluorescence image guided optical coherence tomography (FG-OCT). We examined its ability to enhance specificity and sensitivity for the noninvasive diagnosis of early bladder cancer. MATERIALS AND METHODS: Transitional cell carcinoma was developed in 54 Fisher 344 female rats by intravesical methyl-nitroso-urea instillations. Two or three rats were diagnosed sequentially by 5-ALA (5-aminolevulinic acid hydrochloride) induced fluorescence imaging, cross-sectional OCT and histological microscopy weekly during weeks 11 to 33 following initial methyl-nitroso-urea instillation to track the course of carcinogenesis. RESULTS: The specificity of fluorescence detection was significantly enhanced by FG-OCT (53% and 93%, respectively, p <0.0001). The sensitivity of fluorescence detection and FG-OCT was 79% and 100%, respectively. CONCLUSIONS: FG-OCT cystoscopy has the potential to diagnose early bladder cancer with high sensitivity and specificity with drastically decreased imaging time compared to that of white light guided OCT cystoscopy.

Phage display selection of peptides that affect prostate carcinoma cells attachment and invasion.

BACKGROUND: Prostate cancer-specific proteins must be identified to serve as diagnostic and prognostic markers. Cell surface proteins are especially important, because they have potential utility as diagnostic markers and therapeutic targets. Identification of ligands for these proteins will allow use of these ligands as diagnostic and therapeutic tools and permit the investigation of receptor function. We performed a search for peptide ligands to prostate cancer cell-specific receptors. METHODS: Peptide phage display library was used to isolate specific ligands to LNCaP prostate carcinoma cells receptors. Selected phage and cognate peptides were investigated for their cancer-related functions, such as the ability to interfere with cell adhesion, spreading, motility, and invasion. RESULTS: Phage designated pg35, blocked spreading of LNCaP cells and their derivatives C4-2 and C4-2b. Cognate peptide did not inhibit spreading, but incubation of C4-2 and C4-2b cells with cognate peptide increased their affinity for endothelial cells and invasiveness. In addition, the peptide activates matrix metalloproteinase (MMP)-2 and 9 in C4-2 and C4-2b cells. CONCLUSIONS: These results indicate that identified ligands may play a role in tumorigenicity and metastatic transformation of prostate cancer. To our knowledge, this is the first identification of a functional cancer-specific peptide ligand using the phage display approach.

Psychological and physical health effects of sexual assaults and nonsexual traumas among male and female United States Army soldiers.

Lifetime trauma history was assessed in a health study of active duty United States Army soldiers. Five hundred fifty-five male and 573 female soldiers in the sample were asked whether they had ever experienced 14 different potentially traumatic experiences, including sexual assaults, violent stressors to self, and terrifying events that occurred to others and were secondarily traumatic through exposure by gaining information or as a witness to the event. Most soldiers had experienced multiple traumas, and premilitary exposure to events was much more common than exposure to events after entering the military. Global measures of current psychological distress and physical health symptoms were predicted by the lifetime number of sexual assaults and traumas to self. Social support from military unit leaders moderated the relationship between accumulated exposure to traumas and both health measures, whereas unit cohesion was directly associated with fewer mental health problems.

Acute histologic changes in human renal tumors after cryoablation.

BACKGROUND AND PURPOSE: Cryoablation is a treatment option for some patients with small, exophytic lesions of the kidney. Several investigators have evaluated the effects of cryoablation in normal renal tissue of animals. The purpose of this study was to investigate the tissue changes following cryoablation in human renal tumors. PATIENTS AND METHODS: We prospectively evaluated patients with solid renal lesions (1.5-1.8 cm) confirmed by CT, MRI, or both. Metastatic work-up for all patients was negative. All lesions were biopsied prior to freezing. Two patients with bilateral renal tumors underwent argon-gas-based CRYOcare System (Endocare, Irvine, CA) treatment via an open approach. A 3-mm cryoprobe was placed directly into each tumor. A single 15-minute freeze preceded an active thaw (helium gas) for each lesion. Iceball dimensions were monitored by intraoperative ultrasonography. After successful cryoablation, partial nephrectomy was performed to remove the lesion, and the renal tissue underwent histologic evaluation. RESULTS: The cryoprobes achieved a temperature of -135 degrees C. No bleeding was noted, and there were no intraoperative or postoperative complications with a mean follow-up of 3 months. Histologically, freezing of renal tissue resulted in coagulative necrosis and hemorrhage beyond the boundaries of the lesions. There was a zone of demarcation between the viable and nonviable tissue. CONCLUSIONS: In our series, cryoablation was effective in destroying tumor tissue in vivo in human kidneys. Freezing was sufficient to achieve a negative surgical margin. Cryoablation of renal tumor is an alternative to the currently available nephron-sparing surgical techniques. The long-term effect of tumor tissue destruction by cryosurgery requires further investigation.

Prevalence and timing of sexual assaults in a sample of male and female U.S. Army soldiers.

OBJECTIVE: This study was designed to assess the prevalence and timing of sexual assault experiences in a sample of U.S. Army soldiers. METHODS: Self-administered surveys were completed by 555 male and 573 female soldiers in combat service and combat service support units. RESULTS: One-fifth of the women reported a completed rape (22.6%), and 50.9% of women and 6.7% of men reported any sexual assault. The majority of sexual assaults occurred before the soldiers entered the military, and 25% of women and 1% of men reported an attempted or completed rape during childhood. Sexual assault history also varied by sociodemographic characteristics. CONCLUSION: Results suggest that a history of childhood sexual abuse may be more widespread among female soldiers than among civilian women, and that ascribed and achieved status characteristics may differentially expose soldiers to sexual assaults both before and after they enter the military. Health care assessments should include details of a soldier's sexual assault history.

Cyclosporin A specifically inhibits function of nuclear proteins involved in T cell activation.

One action of cyclosporin A thought to be central to many of its immunosuppressive effects is its ability to inhibit the early events of T lymphocyte activation such as lymphokine gene transcription in response to signals initiated at the antigen receptor. Cyclosporin A was found to specifically inhibit the appearance of DNA binding activity of NF-AT, AP-3, and to a lesser extent NF-kappa B, nuclear proteins that appear to be important in the transcriptional activation of the genes for interleukin-2 and its receptor, as well as several other lymphokines. In addition, cyclosporin A abolished the ability of the NF-AT binding site to activate a linked promoter in transfected mitogen-stimulated T lymphocytes and in lymphocytes from transgenic mice. These results indicate that cyclosporin A either directly inhibits the function of nuclear proteins critical to T lymphocyte activation or inhibits the action of a more proximal member of the signal transmission cascade leading from the antigen receptor to the nucleus.

Identification of a putative regulator of early T cell activation genes.

Molecules involved in the antigen receptor-dependent regulation of early T cell activation genes were investigated with the use of functional sequences of the T cell activation-specific enhancer of interleukin-2 (IL-2). One of these sequences forms a protein complex, NFAT-1, specifically with nuclear extracts of activated T cells. This complex appeared 10 to 25 minutes before the activation of the IL-2 gene. Studies with inhibitors of protein synthesis indicated that the time of synthesis of the activator of the IL-2 gene in Jurkat T cells corresponds to the time of appearance of NFAT-1. NFAT-1, or a very similar protein, bound functional sequences of the long terminal repeat (LTR) of the human immunodeficiency virus type 1; the LTR of this virus is known to be stimulated during early T cell activation. The binding site for this complex activated a linked promoter after transfection into antigen receptor-activated T cells but not other cell types. These characteristics suggest that NFAT-1 transmits signals initiated at the T cell antigen receptor.

Characterization of antigen receptor response elements within the interleukin-2 enhancer.

T-cell activation and induction of interleukin-2 (IL-2) expression in human T lymphocytes require both interaction of foreign antigen with the T-cell antigen receptor and protein kinase C (PKC) stimulation. Agents such as phorbol 12-myristate 13-acetate (PMA) that stimulate PKC augment the effects of antigen but are not sufficient for IL-2 activation. By analysis of deletion mutants, we identified three DNA sequences extending from -73 to -89, -217 to -255, and -263 to -279, designated IL-2 sites A, D, and E, respectively, that are required for maximal induction of IL-2 expression. One of these regions, site E, interacted with a protein (NF-IL-2E) present only in the nuclei of cells which have been stimulated. The other two sequences interacted with a protein (NF-IL-2A) that is constitutively expressed in T cells. When multiple tandem copies of either the E site or the A site were placed upstream of the gamma-fibrinogen promoter, they activated expression via this promoter in response to signals initiated at the antigen receptor but not following PMA stimulation. For this reason, we denoted them antigen receptor response elements. The uncoupling of antigen receptor and PKC requirements in these studies indicates that these signal pathways are, at least in part, distinct and integrated at the level of the gene.

A 275 basepair fragment at the 5' end of the interleukin 2 gene enhances expression from a heterologous promoter in response to signals from the T cell antigen receptor.

Using a transient transfection assay, we have defined the sequences required for the activation of the IL-2 gene in response to signals from the T cell antigen receptor. To do so we have transfected the human T cell line Jurkat with hybrid DNA constructs in which fragments from the IL-2 gene are linked to an indicator gene. The indicator gene product, as well as IL-2 production from the endogenous IL-2 gene were assayed after activation of the transfected Jurkat cells by various stimuli. We have demonstrated that a 275 bp fragment stretching from 52 to 326 bp upstream of the IL-2 gene transcription initiation site is required for expression of the linked indicator gene. This IL-2 gene fragment has several of the characteristics of a transcriptional enhancer element, in that it functions in both orientations and will enhance the expression from the promoter of an unrelated gene. Such enhancement occurred only after activation of Jurkat cells through the T cell antigen receptor. More specifically, this 275 bp fragment activated the expression of a linked gene after binding of a monoclonal antibody to the Jurkat T cell antigen receptor in the presence of PMA. In addition, calcium ionophore, which circumvents antigen receptor binding in T cell activation, induced the expression of the linked gene through this 275 bp sequence, in the presence of PMA. Finally, in a mutant Jurkat cell line lacking T3/antigen receptor complexes at the cell surface, no expression due to the IL-2 5' flanking region was seen after exposure to antibody to the T cell antigen receptor plus PMA or to PHA plus PMA. In contrast, calcium ionophore plus PMA did induce the expression of a linked gene through the IL-2 5' flanking region in the mutant Jurkat cell line. The responsiveness of the transfected hybrid genes containing the IL-2 regulatory region paralleled the expression of the endogenous IL-2 gene, as determined by IL-2 bioassays. We conclude that the 275 bp IL-2 sequence (-326 to -52 bp) is a target for the signal pathway originating at the T cell antigen receptor. Definition of this 275 bp target sequence should now permit the isolation of the molecules that bind to and activate the IL-2 gene.

Retroviral activation of interleukin 2 gene in a gibbon ape T cell lymphoma line.

The gibbon ape leukemia virus (GaLVSF)-infected T cell line, MLA 144, was established from the lymphoma of a gibbon ape. The cell line constitutively expresses IL-2 and its receptor, implying that an autocrine mechanism could be responsible for or contribute toward its growth. To explore the mechanism of constitutive IL-2 expression in MLA 144, we have isolated and characterized cosmid clones representing a normal and a doubly inserted IL-2 allele in this cell line. The map of the normal MLA 144 IL-2 allele closely resembles that of the normal human IL-2 gene. The abnormal allele contains a 3' insertion that is a GaLVSF provirus with two long terminal repeats (LTR) and an internal 3.25 kb deletion. At the 5' end of the abnormal allele is a second insertion that DNA sequencing showed to be an isolated GaLVSF LTR with a transcriptional orientation opposing that of the IL-2 gene. We demonstrate by Northern blotting analysis that the vast majority of transcripts are from the abnormal allele, implying that one or both retroviral insertions are responsible for constitutive expression of the allele.

Promoter region of interleukin-2 gene undergoes chromatin structure changes and confers inducibility on chloramphenicol acetyltransferase gene during activation of T cells.

The chromatin structure of the interleukin-2 (IL-2) gene was probed by DNase I treatment of isolated nuclei. The 5' region of the IL-2 gene contains three regions of hypersensitivity to DNase I. When peripheral blood T cells or Jurkat T cells are stimulated with mitogens, IL-2 message is induced, and the promoter region of the IL-2 gene develops an additional hypersensitive site. This suggests that a DNA sequence close to the transcriptional start site is involved in the transduction of the extracellular signal. Such a conclusion is further supported by DNA transfection experiments. A short segment of DNA, which includes the region of induced hypersensitivity, confers inducibility on the linked chloramphenicol acetyltransferase gene in transiently transfected Jurkat cells. In addition, cells of nonhematopoietic origins exhibit a strikingly different chromatin pattern of IL-2, suggesting a role during differentiation for some of the hypersensitive sites.