Photophysics of pi-conjugated metal-organic oligomers: aryleneethynylenes that contain the (bpy)Re(CO)(3)Cl chromophore.

A comprehensive study of a series of four monodisperse, metal-organic pi-conjugated oligomers of varying length is reported. The oligomers are based on the aryleneethynylene architecture, and they contain a 2,2'-bipyridine-5,5'-diyl (bpy) metal binding unit. The photophysical properties of the free oligomers and their complexes with the (L)Re(I)(CO)(3)X chromophore (where L = the bpy-oligomer and X = Cl or NCCH(3)) were explored by a variety of methods including electrochemistry, UV-visible absorption, variable temperature photoluminescence (PL), transient absorption (TA), and time-resolved electron paramagnetic spectroscopy (TREPR). The absorption of the free oligomers and the metal complexes is dominated by the pi,pi* transitions of the pi-conjugated oligomers. The free oligomers feature a strong blue fluorescence that is quenched entirely in the (L)Re(I)(CO)(3)X complexes. The metal-oligomers feature a weak, relatively long-lived red photoluminescence that is assigned to emission from both the (3)pi,pi* manifold of the pi-conjugated system and the dpi Re --> pi* bpy-oligomer metal-to-ligand charge transfer ((3)MLCT) state. On the basis of a detailed analysis of the PL, TA, and TREPR results an excited-state model is developed which indicates that the oligomer-based (3)pi,pi* state and the (3)MLCT states are in close energetic proximity. Consequently the photophysical properties reflect a composite of the properties of the two excited-state manifolds.

Photophysics of phenyleneethynylene metal-organic oligomers. Probing the lowest excited state by time-resolved IR spectroscopy.

The long-lived excited state in a series of metal-organic phenyleneethynylene oligomers is probed by UV-visible and infrared transient absorption spectroscopy.

In vivo exposure to ultraviolet radiation enhances pathogenic effects of murine leukemia virus, LP-BM5, in murine acquired immunodeficiency syndrome.

LP-BM5 murine leukemia virus (MuLV) induces an immunodeficiency syndrome (MAIDS) in C57BL/6 mice which resembles immunological abnormalities observed in early stages of human AIDS. In our study, MAIDS virus-infected mice were exposed to low doses of ultraviolet radiation (UVR) before and after virus inoculation and compared with MAIDS-infected but not UVR-exposed mice. In all tested parameters (blood IgM levels; mitogenic responses to PHA, ConA, LPS and anti-mu; MLR; antigenic response to SRBC; enlargement and histopathologic changes of the spleen) we observed the same trend: changes due to MAIDS infection were more pronounced in the UVR-exposed group than in the unexposed group. Statistically significant differences between these two groups were seen for mitogenic responses at two different time points after virus inoculation. These results demonstrate that in vivo UVR exposure enhances the immunosuppressive effects of a retroviral infection. UVR exposure may affect the progression of AIDS in a similar manner.

UVC modulation of epidermal growth factor receptor number in HeLa S3 cells.

Induction of transforming growth factor alpha (TGF alpha) in human cell lines by 254 nm ultraviolet radiation (UVC) suggests that TGF alpha may have an autocrine role in UV-induced tumorigenesis. Binding of TGF alpha to epidermal growth factor receptor (EGFR) is an important initial step in transducing the signal for cell division. Experiments reported herein were designed to determine whether, in addition to inducing TGF alpha, UVC might also induce changes in the levels of EGFR on HeLa S3 cells [125I]EGF binding to HeLa S3 cells was inhibited 8 h after exposure to 7 J/m2 UVC radiation followed by increased [125I]EGF binding 16-32 h after irradiation. Scatchard analysis of EGF binding at 28 h indicated that irradiated cells had 60% more receptors with no differences in apparent binding affinities (56,300 +/- 5494 receptors versus 34,900 +/- 1899 receptors in sham-irradiated cells). Cell cycle analysis at 8 h post-UVC indicated that cells had slowed traverse of S-phase, but by 24 and 48 h, times at which increases in [125I]EGF were evident, cell cycle distributions were essentially back to normal. These results indicate that UVC modulates EGFR numbers in HeLa S3 cells and suggest that solar radiation may modulate EGFR numbers in keratinocytes or other cells in the skin. The presence of UV-induced growth factors such as TGF alpha and increased levels of EGFR may result in sustained cell proliferation by autocrine or paracrine mechanisms. These populations of cycling cells would then be at risk for subsequent mutational events that result in transformation to a tumorigenic state.

Sunscreen protection against UV-induced pyrimidine dimers in DNA of human skin in situ.

We have determined the ability of a chemical sunscreen with a sun protection factor (SPF) of 15 to protect human skin from ultraviolet (UV) radiation-induced DNA damage. The DNA damage was susceptible to cleavage by Micrococcus luteus UV endonuclease, which recognizes pyrimidine dimers in DNA. An alkaline agarose gel electrophoresis method was used to quantify the number of pyrimidine dimers in nonradioactive DNA from skin biopsies of 5 individuals irradiated with UV from a solar simulator. After exposure to an equivalent dose of UV, the number of pyrimidine dimers was 0.8 per 10(7) bases in sunscreen-treated skin as compared with 32 dimers per 10(7) bases in untreated skin. This assay provides a means of determining the efficacy of sunscreens in protecting skin from UV-induced DNA damage.

Relationship between circulating interferon and anti-interferon antibodies and impaired natural killer cell activity in systemic lupus erythematosus.

Interferon (IFN) production and response are impaired in a high percentage of systemic lupus erythematosus (SLE) patients. In addition, elevated serum levels of alpha-IFN or anti-alpha-IFN antibodies are present in some SLE patients. This study examined the relationship of circulating IFN and anti-IFN antibodies to the impairment of natural killer (NK) cell function in SLE. All 15 SLE patients studied had measurable circulating alpha-IFN, while the normal controls had minimal serum IFN. Neither patient nor control sera contained any detectable anti-alpha-IFN activity. However, most of the SLE patients demonstrated defects in NK cell function. Because these defects in NK cell function appeared to be associated with circulating IFN, but not anti-IFN, antibodies, the effect of prolonged in vitro IFN exposure on NK cell function of peripheral blood mononuclear cells was determined. It was found that prolonged exposure to IFN induced both an apparent defect in IFN response and a definite impairment of baseline NK cell function. These results suggest that prolonged elevation of circulating alpha-IFN levels could be responsible, in part, for the defects in natural cytotoxicity present in SLE.

Lymphokines in autoimmunity: relationship between interleukin-2 and interferon-gamma production in systemic lupus erythematosus.

Interleukin-2 (IL-2) and interferon (IFN) are potent immunoregulatory factors. Recently, it has been demonstrated that IL-2 production is necessary for IFN-gamma synthesis. Thus, defects in IL-2 production could lead to defects in IFN-gamma production. Indeed, in systemic lupus erythematosus, defects in IFN-gamma and IL-2 production have been noted. To test this hypothesis, IL-2 and IFN-gamma production rates by peripheral blood mononuclear cells PBMC were measured simultaneously after stimulation by concanavalin A (Con A) and phorbol myristic acetate (PMA). IL-2 activity was determined employing HT-2, a cell line with an absolute growth requirement for IL-2. IFN-gamma activity was assessed using a standard viral plaque inhibition assay. IL-2 production in normal controls (n = 6) was 14.7 +/- 9.4 units/ml and in SLE patients (n = 10), 18.0 +/- 10.5 units/ml (P greater than 0.05). IFN-gamma production in controls was 74.7 +/- 43.7 units/ml and in SLE patients, 68.8 +/- 35.4 units/ml (P greater than 0.05). Only one SLE patient, who had moderately active disease, demonstrated defects in IL-2 synthesis (less than 4 units/ml) and IFN-gamma production (16 units/ml). Thus, production of these lymphokines in SLE patients was largely normal in response to Con A-PMA. These results imply that the intrinsic pathways of IFN and IL-2 production are basically normal in most SLE patients although defects in production of these lymphokines can occur. The defects in IL-2 and IFN production in SLE previously reported may be secondary to an impaired cellular response to the selected inducing agent rather than a primary defect in the actual ability to produce these important lymphokines.

Mitochondrial protein synthesis in Chinese hamster cells (line CHO) synchronized by isoleucine deprivation.

Mitochondrial protein synthesis was measured in line CHO cells after phases of the cell cycle were synchronized by isoleucine deprivation or mitotic selection. Maximum incorporation of [3H] leucine into mitochondrial polypeptides occurred within 2 hours after isoleucine was added to initiate G1 traverse. In cells synchronized in G1 by mitotic selection, the rate of mitochondrial protein synthesis was fairly constant throughout the cell cycle. SDS-polyacrylamide gel electrophoretic profiles of labeled mitochondrial polypeptides were similar in cells synchronized by either isoleucine deprivation or mitotic selection. Obvious changes in the distribution of polypeptides were not detected during various phases of the cell cycle. The increased rate of incorporation of [3H] leucine into mitochondrial polypeptides after reversal of G1-arrest may indicate that mitochondrial protein synthesis and possibly mitochondrial biogenesis are synchronized in CHO cells deprived of isoleucine.

Propionate-induced synthesis of odd-chain-length fatty acids by Escherichia coli.

Exogenous propionate is incorporated in vivo by Escherichia coli as a primer to produce lipids with fatty acids of odd chain lengths. This provides a method for the specific labeling of the three terminal carbons in the fatty acyl chains of phospholipids.

Effect of isoleucine deprivation of rabbitpox virus DNA synthesis in mouse L cells.

Synthesis of rabbitpox DNA was inhibited in mouse L cells deprived of isoleucine. Time-course patterns of incorporation of radiolabelled precursors into viral DNA revealed that synthesis of viral DNA began about 6 h after reversal of the isoleucine-deficient state.

Isolation and characterization of a herpesvirus from wild turkeys (Meleagris gallopavo osceola) in Florida.

Viral agents producing both a syncytial-type cytopathic effect and type A intranuclear inclusion bodies in vitro were isolated from the kidneys of five of 10 wild turkeys. A plaque assay system for viral infectivity was developed and used to characterize one of the wild turkey viruses (WTV). WTV replication was inhibited by 5-bromodeoxyuridine, indicating the virus contained DNA as its genetic material. Virus infectivity could be transferred only as viable whole cell preparations; one cycle of rapid freezing and thawing completely inactivated the virus. Typical herpes-like virions were found within the nuclei when cells infected with WTV were examined by electron microscopy. WTV had characteristics typical of the herpes group of viruses.

Detection of G1 proteins in Chinese hamster cells synchronized by isoleucine deprivation or mitotic selection.

Examination of labeling patterns of proteins in Chinese hamster cells(line CHO) revealed the presence of a class of protein(s) that is synthesized during G1 phase of the cell cycle. Cells arrested in G1 by isoleucine (Ile) deprivation were prelabeded with [14-C]Ile, induced to traverse G1 by addition of unlabeled Ile, and labeled with [3-H]Ile at hourly intervals. Cells were fractionated into neclear and cytoplasmic portions, and proteins were separated by sodium dodecyl sulfate-polyacrylamide get electrophoresis. Gel profiles of proteins in the 45,000-160,000 mol wt range from the cytoplasm of cells in G1 were similar to those from cells arrested in G1 except for the presence of a mojor peak of [1-H]Ile incorporated into a protein(s) of approximately 80,000 mol wt. Peaks of net [3-H]Ile incorporation were not detected in neclear preparations. Cellular fractionation by differential centrifugation showed the peak I protein was located in the soluble supernatant fraction of the cytoplasm. Time-course studies showed that synthesis of this protein began 1-2 h after initiation of G1 traverse; the protein reached maximum levels in 4-6 h and was reduced to undetectable levels by 9 h. A cytoplasmic protein with similar electrophoretic mobility was found in G1 phase of cells synchronized by mitotic selection. This class of proteins is synthesized by cells before entry into S phase and may be involved in initiation of DNA synthesis.

An accurate slicer of disc acrylamide gels devised by simple modification of a density gradient fractionator.

A commercially available density gradient fractionator has been modified in a simple and inexpensive way to yield a device for accurately slicing disc acrylamide gels. Attractive features of this device include convenience of operation and the capacity to easily vary thickness of the gel slice.

Synchronization of mitochondrial DNA synthesis in Chinese hamster cells (line CHO) deprived of isoleucine.

Mitochondrial DNA (mit-DNA) synthesis was compared in suspension cultures of Chinese hamster cells (line CHO) whose cell cycle events had been synchronized by isoleucine deprivation or mitotic selection. At hourly intervals during cell cycle progression, synchronized cells were exposed to tritiated thymidine ([(3)H]TdR), homogenized, and nuclei and mitochondria isolated by differential centrifugation. Mit-DNA and nuclear DNA were isolated and incorporation of radioisotope measured as counts per minute ([(3)H]TdR) per microgram DNA. Mit-DNA synthesis in cells synchronized by mitotic selection began after 4 h and continued for approximately 9 h. This time-course pattern resembled that of nuclear DNA synthesis. In contrast, mit-DNA synthesis in cells synchronized by isoleucine deprivation did not begin until 9-12 h after addition of isoleucine and virtually all [(3)H]TdR was incorporated during a 3-h interval. We have concluded from these results that mit-DNA synthesis is inhibited in CHO cells which are arrested in G(1) because of isoleucine deprivation and that addition of isoleucine stimulates synchronous synthesis of mit-DNA. We believe this method of synchronizing mit-DNA synthesis may be of value in studies of factors which regulate synthesis of mit-DNA.

Regulation of initiation of DNA synthesis in Chinese hamster cells. I. Production of stable, reversible G1-arrested populations in suspension culture.

Suspension cultures of Chinese hamster cells (line CHO) were grown to stationary phase (approximately 8-9 x 10(5) cells/ml) in F-10 medium. Cells remained viable (95%) for at least 80 hr in stationary phase, and essentially all of the cells were in G(1) Upon resuspension or dilution with fresh medium, the cells were induced to resume traverse of the life cycle in in synchrony, and the patterns of DNA synthesis and division were similar to those observed in cultures prepared by mitotic selection. Immediately after dilution, the rates of synthesis of RNA and protein increased threefold. This system provides a simple technique for production of large quantities of highly synchronized cells and may ultimately provide information on the biochemical mechanisms regulating cell-cycle traverse.

Cell-associated nature of cottontail rabbit herpesvirus in vitro.

The cottontail rabbit herpesvirus is presently classified in subgroup A of the herpesviruses, whereas the virus is strongly cell associated and belongs in subgroup B.