Dielectric tensor of tetracene single crystals: the effect of anisotropy on polarized absorption and emission spectra.

The full UV-visible dielectric tensor and the corresponding directions of the principal axes of triclinic tetracene crystals are reported as deduced either by polarized absorption and ellipsometry measurements or by calculations based on the molecular and crystallographic data. The results allow the attribution of the polarized bands observed in both absorption and photoluminescence emission spectra. In particular, the spectral line shape and polarization of the emission are found to depend on the sample thickness, and the effect is attributed to the modification of the state of polarization of the emitted light during its propagation inside the crystal. Indeed, the directions of polarization of the lowest optical transitions and the directions of the principal axes of the dielectric tensor are demonstrated not to coincide, in contrast to the assumptions typically made in the literature, thus causing the mixed transverse/longitudinal character of light propagation.

Reclassifying exciton-phonon coupling in molecular aggregates: evidence of strong nonadiabatic coupling in oligothiophene crystals.

Exciton-phonon (EP) coupling in molecular aggregates is reexamined in cases where extended intermolecular interactions result in low-energy excitons with high effective masses. The analysis is based on a single intramolecular vibrational mode with frequency omega0 and Huang-Rhys factor lambda2. When the curvature Jc at the exciton band bottom is much smaller than the free-exciton Davydov splitting W, the strength of the EP coupling is determined by comparing the nuclear relaxation energy lambda2omega0 with the curvature. In this way, weak (lambda2omega0<<4piJc), intermediate I (lambda2omega0 approximately 4piJc), and strong I (lambda2omega0>>4piJc) coupling regimes are introduced. The conventional intermediate (lambda2omega0 approximately W) and strong (lambda2omega0>>W) EP coupling regimes originally defined by Simpson and Peterson [J. Chem. Phys. 26, 588 (1957)] are based solely on the Davydov splitting and are referred to here as intermediate II and strong II regimes, respectively. Within the intermediate I and strong I regimes the near degeneracy of the low-energy excitons allows efficient nonadiabatic coupling, resulting in a spectral splitting between the b- and ac-polarized first replicas in the vibronic progression characterizing optical absorption. Such spectral signatures are clearly observed in OT4 thin films and crystals, where splittings for the lowest energy mode with omega0=161 cm(-1) are as large as 30 cm(-1) with a small variation due to sample disorder. Numerical calculations using a multiphonon BO basis set and a Hamiltonian including linear EP coupling yield excellent agreement with experiment.

Exciton self-trapping in tetrafluoro-dimethyl-aminoacridine single crystals.

The UV-visible optical spectra of 1,2,3,4-tetrafluoro-7-(N,N)dimethyl-amino-acridine single crystals are reported. The results are discussed on the basis of the molecular transitions and crystal packing in the framework of the theory of molecular excitons under a fluctuating potential field due to dynamic disorder. A strong local geometry distortion is demonstrated by applying the Urbach rule to the absorption tails, which is the amplitude of the local potential fluctuation being larger than the intermolecular transfer energy. The lineshape and linewidth of the emission band and its temperature dependence give further evidence of exciton self-trapping.

Effect of static and dynamic disorder on exciton mobility in oligothiophenes.

The polarized optical absorption spectra of different quaterthiophene single crystals in the energy region of the exciton bands originating from the first molecular transition are reported as measured in the temperatures ranging from 7 to 140 K. The intrinsic higher mobility of the b-polarized 0-0 a(u) exciton both with respect to its replicas and to the a-polarized structures is demonstrated in high quality crystals at the lowest temperatures. The influence of structural disorder on mobility is discussed considering, for the different samples, the measured lineshape and linewidth of the absorption peaks, and the relative lineshift and intensity ratio between the 0-0 a(u) line and its first replica at the lowest temperature. The influence of dynamic disorder is discussed considering the lineshape and linewidth of the measured peaks as a function of temperature for both polarizations in the framework of the exciton-phonon coupling theory.

Measured Davydov splitting in oligothiophene crystals.

The polarized absorption spectra of single crystals of oligothiophenes in a wide spectral range are reported. The experimental procedure is discussed, underlying several details which are relevant to obtain reliable spectra particularly for samples of increasing thickness. On the basis of these considerations, it has been possible to fully detect the transition to the upper Davydov exciton originating from the first molecular state. The position and shape of the main exciton peak in these materials are compared and discussed, taking into consideration the molecular arrangement and the longitudinal contribution which depends on the transition moment orientation. The Davydov splitting values as deduced from the experimental data at room temperature are also reported either for the first vibronic replica or for the electronic transition as a whole. The difference between the purely transverse and the measured Davydov splitting is discussed.

Optical transverse excitation in quaterthiophene crystals by ultraviolet-visible internal and attenuated total reflection.

We report internal and attenuated total reflection of light at the interface between glass and a quaterthiophene crystal in the spectral region of the electronic transitions. The bands corresponding to the absorption of the a(u) and b(u) Frenkel exciton states are detected for different polarization of the incident light. In particular, the wave-normal vector being almost perpendicular to the b(u) transition dipole moment allows its transverse component to be accessed, whose excitation in conventional external reflection or transmission spectroscopies is forbidden.

Directional dispersion in absorbance spectra of oligothiophene crystals.

Due to the large oscillator strength of the first molecular transition in oligothiophenes, a strong directional dispersion of the b(u) exciton transition is expected originating from the macroscopic polarization field. Examining such dispersion unambiguously usually requires different faces to be accessible for the optical measurements. Alternatively, measurements carried out at different angles of incidence are met with intrinsic limits due to the peculiarities of wave propagation in such anisotropic systems. In order to demonstrate these limits along with the experimental difficulties involved, we examine refraction and absorption of light in these crystals and discuss the effects of directional dispersion on the absorbance spectra of quaterthiophene crystals.

Polarized absorption of quaterthiophene single crystals.

Polarized optical spectra of quaterthiophene single crystals are reported over a wide spectral range for different planes and angles of incidence corresponding to either transverse or mixed longitudinal/transverse wave propagation. From the absorbance and reflectance spectra the corresponding absorption coefficients are deduced. In the region of the first molecular transition, polarized exciton states of Frenkel origin are found to be responsible for a strong optical anisotropy and for the modification of the state of polarization during light propagation in the crystal. Transmission measurements with crossed polarizer and analyzer allow to clearly identify Frenkel replica of the principal transition.

Reflectivity and anisotropic optical functions of quaterthiophene single crystals.

Polarized reflectance spectra of quaterthiophene single crystals are reported with different angles and planes of incidence. The strong dependence of the spectral features on the experimental configuration is described by an orthorhombic model and the components of the complex diagonal dielectric tensor are given.

Two phases of HIV RNA decay in CSF during initial days of multidrug therapy.

BACKGROUND: Defining cellular and tissue sources of HIV-1 in CSF is important for understanding disease pathogenesis and optimal therapies for HIV infection in the brain. OBJECTIVE: To identify the time of maximal viral decay in CSF during the initial days of antiretroviral therapy. METHODS: Serial CSF and plasma data were available from four adults who underwent ultraintensive CSF sampling for 48 hours at baseline and again beginning 72 hours after starting antiretroviral therapy. Regression lines were generated using HIV-1 RNA data from 17 on-treatment serial CSF samples obtained at 3-hour intervals. Viral RNA was quantified by Nuclisens and Amplicor HIV-1 Monitor assays. RESULTS: Extrapolation of regression lines intersected baseline below actual baseline CSF HIV-1 RNA concentrations, indicating that virus decayed most rapidly on days 1 through 3 with half-lives of no more than 0.9 to 2.8 days. Half-lives on days 4 and 5 ranged from 1.3 to 4.9 days. Plasma data also showed early rapid decay. CONCLUSIONS: Multiple phases of viral decay suggest that virus in CSF originates from at least two sources during untreated, asymptomatic HIV-1 infection. The short half-life indicates that the primary source is CD4+ T cells. Sampling during days 1 through 3 and different stages of disease will better define sources of virus.

Intrinsic excitonic luminescence in odd and even numbered oligothiophenes.

The emission properties of crystalline oligothiophenes (OTs) are investigated and related to the different selection rules arising from the number of rings. In odd OT crystals the purely electronic emission is absent since the molecular A1-1B(1) transition dipoles cancel at the bottom of the excitonic band. On the contrary, in crystals of even OTs this transition is allowed due to the constructive interference of the off-axis components of the molecular transition dipole. It possesses a polarization in the herringbone plane and exhibits superradiant behavior.

Comparison of Roche MONITOR and Organon Teknika NucliSens assays to quantify human immunodeficiency virus type 1 RNA in cerebrospinal fluid.

We compared Roche MONITOR and Organon Teknika NucliSens assays for human immunodeficiency virus type 1 (HIV-1) RNA in cerebrospinal fluid (CSF). Results of 282 assays were highly correlated (r = 0.826), with MONITOR values being 0.29 +/- 0.4 log(10) copies/ml (mean +/- standard deviation) values. Both assays can reliably quantify HIV-1 RNA in CSF.

QS-21 promotes an adjuvant effect allowing for reduced antigen dose during HIV-1 envelope subunit immunization in humans.

Three separate studies were undertaken in HIV-1 uninfected persons to determine if the adjuvant QS-21 improves the magnitude or kinetics of immune responses induced by recombinant soluble gp120 HIV-1(MN) protein (rsgp120) immunization. The QS-21 was administered at two doses (50 and 100 microg), either alone or in combination with aluminum hydroxide (600 microg). At the highest doses of rsgp120 (100, 300, and 600 microg), QS-21 exerted no significant effect on either binding or neutralizing antibody titers. Antibody binding and neutralizing responses fell dramatically when rsgp120, formulated with alum alone, was given at low doses (3 and 30 microg). In contrast, antibody responses similar in titer to those in the high dose antigen groups were induced with the low dose rsgp120 formulated with QS-21. In addition, the lymphocyte proliferation and delayed type hypersensitivity skin testing were superior in the QS-21 recipients compared with the alum recipients at the low antigen doses. Moderate to severe pain was observed in majority of the volunteers receiving QS-21 formulations, and vasovagal episodes and hypertension were not infrequent. Thus, the use of QS-21 may provide a means to reduce the dose of a soluble protein immunogen.

Expression of vaccinia E3L and K3L genes by a novel recombinant canarypox HIV vaccine vector enhances HIV-1 pseudovirion production and inhibits apoptosis in human cells.

Poxviruses that are attenuated for growth in human cells provide a safe means of HIV antigen expression and are capable of eliciting HIV-specific immune responses, including CD8+ cytotoxic T-lymphocyte (CTL) responses. HIV-1 antigen expression in human cells by attenuated poxvirus vectors may be limited by interferon-mediated host defense mechanisms. To enhance HIV antigen expression in human cells, the vaccinia virus E3L and K3L genes were inserted into a canarypox vector that expresses HIV-1 Gag, Env, and a Nef/Pol polyepitope string. E3L and K3L markedly reduced the activation of the double-stranded RNA-dependent protein kinase, PKR, and led to a significant reduction in apoptosis in HeLa cells. Production and release of HIV-1 antigen in the form of pseudovirions was enhanced in both duration and magnitude by this vector modification. The addition of immunomodulatory genes to attenuated poxviruses represents a novel strategy for enhancing antigen production by live vector HIV vaccine candidates.

Tracking the assembly pathway of human immunodeficiency virus type 1 Gag deletion mutants by immunogold labeling.

The Pr55gag gene product of human immunodeficiency virus type 1 (HIV-1) is sufficient to direct the formation of retrovirus-like particles (RVLPs). Recent biochemical evidence has indicated the presence of Gag intermediates in the cytoplasm; however, the Gag assembly process into RVLPs remains incompletely defined. The authors present here the subcellular localization of Gag mutant proteins in BSC40 and Jurkat cells by immunoelectron microscopy (IEM). The full Gag/Pol and Gag precursors, a C-terminal deletion mutant lacking a portion of nucleocapsid (NC), and all p6Gag gave rise to similar levels of RVLPs at the cell surface. A C-terminal deletion of all NC and p6Gag abrogated particle formation, whereas p24 was found in patches at the cell surface. Deletion of matrix (MA) sequences from Gag resulted in intracellular particles, and myristylation was not required for particle formation in the context of the MA deletion. Matrix expression was enhanced with Gag/Pol or Env coexpression as determined by semiquantitative IEM. p24 protein was targeted at vacuolar and mitochondrial membranes, but not at Golgi cisternae. In addition, aggregations of Gag intermediates and RVLPs in the cytoplasm, rough endoplasmic reticulum, cisternae, and mitochondria were noted. These results provide defined in situ evidence that HIV-1 particle assembly occurs in the cytosol in addition to budding at most intracellular membranes.

Comparison of human immunodeficiency virus type 1 RNA sequence heterogeneity in cerebrospinal fluid and plasma.

The source of human immunodeficiency virus type 1 (HIV-1) RNA in cerebrospinal fluid (CSF) during HIV-1 infection is uncertain. The sequence heterogeneity of HIV-1 RNA in simultaneous CSF and plasma samples was characterized for five patients at the baseline and during the first week of antiretroviral therapy by two commercial genotyping methodologies. In individual subjects, the sequences in CSF samples differed significantly from those in plasma. In contrast, the viral sequences in CSF at the baseline did not differ from the sequences in CSF during treatment. Similarly, viral sequences in plasma did not vary over this interval. This study provides evidence that HIV-1 RNA in CSF and plasma arise from distinct compartments.

Steady-state pharmacokinetics of indinavir in cerebrospinal fluid and plasma among adults with human immunodeficiency virus type 1 infection.

To characterize steady-state indinavir pharmacokinetics in cerebrospinal fluid and plasma, 8 adults infected with human immunodeficiency virus underwent intensive cerebrospinal fluid sampling while receiving indinavir (800 mg every 8 hours) plus nucleoside reverse transcriptase inhibitors. Nine and 11 serial cerebrospinal fluid and plasma samples, respectively, were obtained from each subject. Free indinavir accounted for 94.3% of the drug in cerebrospinal fluid and 41.7% in plasma. Mean values of cerebrospinal fluid peak concentration, concentration at 8 hours, and area under the concentration-time profile calculated over the interval 0 to 8 hours [AUC(0-8)] for free indinavir were 294 nmol/L, 122 nmol/L, and 1616 nmol/L x h, respectively. The cerebrospinal fluid-to-plasma AUC(0-8) ratio for free indinavir was 14.7% +/- 2.6% and did not correlate with indexes of blood-brain barrier integrity or intrathecal immune activation. Indinavir achieves levels in cerebrospinal fluid that should contribute to control of human immunodeficiency virus type 1 replication in this compartment. The cerebrospinal fluid-to-plasma AUC(0-8) ratio suggests clearance mechanisms in addition to passive diffusion across the blood-cerebrospinal fluid barrier, perhaps by P-glycoprotein-mediated efflux.

Evidence of a source of HIV type 1 within the central nervous system by ultraintensive sampling of cerebrospinal fluid and plasma.

Defining the source of HIV-1 RNA in cerebrospinal fluid (CSF) will facilitate studies of treatment efficacy in the brain. Four antiretroviral drug-naive adults underwent two 48-hr ultraintensive CSF sampling procedures, once at baseline and again beginning on day 4 after initiating three-drug therapy with stavudine, lamivudine, and nelfinavir. At baseline, constant CSF HIV-1 RNA concentrations were maintained by daily entry of at least 10(4) to 10(6) HIV-1 RNA copies into CSF. Change from baseline to day 5 ranged from -0.38 to -1.18 log(10) HIV-1 RNA copies/ml in CSF, and from -0.80 to -1.33 log(10) HIV-1 RNA copies/ml in plasma, with no correlation between CSF and plasma changes. There was no evidence of genotypic or phenotypic viral resistance in either CSF or plasma. With regard to pharmacokinetics, mean CSF-to-plasma area-under-the-curve (AUC) ratios were 38.9% for stavudine and 15.3% for lamivudine. Nelfinavir and its active M8 metabolite could not be accurately quantified in CSF, although plasma M8 peak level and AUC(0-8hr) correlated with CSF HIV-1 RNA decline. This study supports the utility of ultraintensive CSF sampling for studying HIV-1 pathogenesis and therapy in the CNS, and provides strong evidence that HIV-1 RNA in CSF arises, at least in part, from a source other than plasma.

Mapping and characterization of the N-terminal I domain of human immunodeficiency virus type 1 Pr55(Gag).

Human immunodeficiency virus (HIV) type 1 particles assemble at the plasma membrane of cells in a manner similar to that of the type C oncoretroviruses. The Pr55(Gag) molecule directs the assembly process and is sufficient for particle assembly in the absence of all other viral gene products. The I domain is an assembly domain that has been previously localized to the nucleocapsid (NC) region of Gag. In this study we utilized a series of Gag-green fluorescent protein (GFP) fusion proteins to precisely identify sequences that constitute the N-terminal I domain of Pr55(Gag). The minimal sequence required for the I domain was localized to the extreme N terminus of NC. Two basic residues (arginine 380 and arginine 384) within the initial seven residues of NC were found to be critical for the function of the N-terminal I domain. The presence of positive charge alone in these two positions, however, was not sufficient to mediate the formation of dense Gag particles. The I domain was required for the formation of detergent-resistant complexes of Gag protein, and confocal microscopy demonstrated that the I domain was also required for the formation of punctate foci of Gag proteins at the plasma membrane. Electron microscopic analysis of cells expressing Gag-GFP fusion constructs with an intact I domain revealed numerous retrovirus-like particles (RVLPs) budding from the plasma membrane, while I domain-deficient constructs failed to generate visible RVLPs. These results provide evidence that Gag-Gag interactions mediated by the I domain play a central role in the assembly of HIV particles.