Photoplethysmographic waveform characteristics of newborns with coarctation of the aorta.

OBJECTIVE: Fetal echocardiography, physical examination and pulse oximetry detect only half of coarctation of aorta (CoA) cases. We aimed to quantify delayed arrival and diminished amplitude of lower extremity photoplethysmographic (PPG) pulses relative to the right hand in affected patients. STUDY DESIGN: We studied 8 CoA infants and 32 healthy controls. The pulse arrival time difference between foot and hand (f-hTD) and pulse amplitude ratio (F/H) were measured on PPG signal waveforms by digitally-determining maxima and minima of systolic decrease of light transmission. Mann-Whitney test was used for group comparisons. RESULTS: In comparison to healthy newborns, CoA infants' PPG waveforms demonstrated prolonged f-hTD (mean±s.d. of 73.2±26.6 versus 35.2±8.3 ms, P<0.001) and lower F/H (0.57±0.26 versus 0.99±0.58, P=0.014). CONCLUSIONS: F-hTD and F/H are quantifiable from hand- and foot-derived PPG waveforms and are significantly different in CoA versus healthy newborns. Larger studies are needed to validate PPG for improved critical congenital heart disease screening.

Imidazoacridinone-dependent lysosomal photodestruction: a pharmacological Trojan horse approach to eradicate multidrug-resistant cancers.

Multidrug resistance (MDR) remains a primary hindrance to curative cancer therapy. Thus, introduction of novel strategies to overcome MDR is of paramount therapeutic significance. Sequestration of chemotherapeutics in lysosomes is an established mechanism of drug resistance. Here, we show that MDR cells display a marked increase in lysosome number. We further demonstrate that imidazoacridinones (IAs), which are cytotoxic fluorochromes, undergo a dramatic compartmentalization in lysosomes because of their hydrophobic weak base nature. We hence developed a novel photoactivation-based pharmacological Trojan horse approach to target and eradicate MDR cancer cells based on photo-rupture of IA-loaded lysosomes and tumor cell lysis via formation of reactive oxygen species. Illumination of IA-loaded cells resulted in lysosomal photodestruction and restoration of parental cell drug sensitivity. Lysosomal photodestruction of MDR cells overexpressing the key MDR efflux transporters ABCG2, ABCB1 or ABCC1 resulted in 10- to 52-fold lower IC(50) values of various IAs, thereby restoring parental cell sensitivity. Finally, in vivo application of this photodynamic therapy strategy after i.v. injection of IAs in human ovarian tumor xenografts in the chorioallantoic membrane model revealed selective destruction of tumors and their associated vasculature. These findings identify lysosomal sequestration of IAs as an Achilles heel of MDR cells that can be harnessed to eradicate MDR tumor cells via lysosomal photodestruction.

A universal epitope-based influenza vaccine and its efficacy against H5N1.

Previous studies have shown that a recombinant vaccine expressing four highly conserved influenza virus epitopes has a potential for a broad spectrum, cross-reactive vaccine; it induced protection against H1, H2 and H3 influenza strains. Here, we report on the evaluation of an epitope-based vaccine in which six conserved epitopes, common to many influenza virus strains are expressed within a recombinant flagellin that serves as both a carrier and adjuvant. In an HLA-A2.1 transgenic mice model, this vaccine induced both humoral and cellular responses and conferred some protection against lethal challenge with the highly pathogenic H5N1 avian influenza strain. Hence, it is expected to protect against future strains as well. The data presented, demonstrate the feasibility of using an array of peptides for vaccination, which might pave the way to an advantageous universal influenza virus vaccine that does not require frequent updates and/or annual immunizations.

Search for correlates of protective immunity conferred by anthrax vaccine.

Vaccination by anthrax protective antigen (PA)-based vaccines requires multiple immunization, underlying the need to develop more efficacious vaccines or alternative vaccination regimens. In spite of the vast use of PA-based vaccines, the definition of a marker for protective immunity is still lacking. Here we describe studies designed to help define such markers. To this end we have immunized guinea pigs by different methods and monitored the immune response and the corresponding extent of protection against a lethal challenge with anthrax spores. Active immunization was performed by a single injection using one of two methods: (i) vaccination with decreasing amounts of PA and (ii) vaccination with constant amounts of PA that had been thermally inactivated for increasing periods. In both studies a direct correlation between survival and neutralizing-antibody titer was found (r(2) = 0.92 and 0.95, respectively). Most significantly, in the two protocols a similar neutralizing-antibody titer range provided 50% protection. Furthermore, in a complementary study involving passive transfer of PA hyperimmune sera to naive animals, a similar correlation between neutralizing-antibody titers and protection was found. In all three immunization studies, neutralization titers of at least 300 were sufficient to confer protection against a dose of 40 50% lethal doses (LD(50)) of virulent anthrax spores of the Vollum strain. Such consistency in the correlation of protective immunity with anti-PA antibody titers was not observed for antibody titers determined by an enzyme-linked immunosorbent assay. Taken together, these results clearly demonstrate that neutralizing antibodies to PA constitute a major component of the protective immunity against anthrax and suggest that this parameter could be used as a surrogate marker for protection.

GroESL proteins facilitate binding of externally added inducer by LuxR protein-containing E. coli cells.

htpR- (rpoH, sigma 32 minus) strain of E. coli harbouring the whole lux system of Vibrio fischeri is very dim. We have recently shown that GroESL proteins fully recover the expression of the lux system in this strain. This work has been undertaken to study our assumption that the GroESL proteins stabilize the LuxR protein, thus enhancing the formation of LuxR-Inducer complex. E. coli htpR- cells harbouring the luxR gene were unable to bind extracellularly added inducer, while late logarithmically growing htpR+ strain bound small quantities of the inducer. Reduction in the nutrient content of the growth medium resulted in a large increase in the capability of these cells to bind the inducer. htpR+ or htpR- E. coli strains harbouring both the luxR and the groESL genes bound large quantities of the inducer. The molecular and ecological significance of these results is discussed.

Formation of the LuxR protein in the Vibrio fischeri lux system is controlled by HtpR through the GroESL proteins.

The transcription of the luminescence (lux) system of Vibrio fischeri is regulated by the LuxR protein and an autoinducer. We previously showed that apart from these regulatory elements, the transcription of the lux system is negatively controlled by the LexA protein and positively controlled by the HtpR protein (sigma 32). This study was conducted in order to elucidate the mode of action of the HtpR protein. Using luxR-lacZ fused genes, we showed that the HtpR protein is essential for the maximum expression of beta-galactosidase activity in Escherichia coli lac mutant cells. Using this construct, we also demonstrated that luxR is preferentially expressed toward the end of the logarithmic phase of growth. Starvation and addition of ethanol significantly advanced the appearance of beta-galactosidase activity in htpR+ cells. The luminescence system of E. coli htpR+ cells harboring the pChv1 plasmid with a deletion in the luxI gene is induced in the presence of low and constant concentrations (150 pg/ml) of the inducer only at a late stage of the logarithmic phase of growth. When the cellular LuxR content is reduced, following 23 generations of exponential growth in Luria broth, a mid-log-phase culture does not respond to the inducer (150 pg/ml). On the basis of the above observations we suggest that the HtpR protein controls the formation of V. fischeri LuxR protein. Preliminary findings indicate that the HtpR protein acts through the chaperonins GroESL. E. coli htpR/pChv1 cells retained their full level of in vivo and in vitro luciferase activities in the presence of multiple copies of groESL genes. The possibility that GroESL proteins stabilize the native form of LuxR protein is discussed.