John A. H. Lee

ABSOLUTE SPECTRAL SENSITIVITY OF PHOTOTUBES AND THE APPLICATION TO THE MEASUREMENT OF THE ABSOLUTE QUANTUM YIELDS OF CHEMILUMINESCENCE AND BIOLUMINESCENCE

Abstract— Methods for the absolute measurement of the spectral sensitivity of phototubes are reviewed in detail. Several experimental arrangements are described for the independent determination of these values to insure the elimination of systematic error. On the basis of these measurements a rapid, simple technique is described for measuring the average absolute spectral sensitivity of a phototube over a defined emission spectrum, thus obviating the original tedious point‐by‐point methods and allowing frequent re‐Standardization. The application of these standardized phototubes to the measurement of the absolute quantum yields of the chemiluminescence of luminol and the bioluminescence of luminous bacteria and the firefly is described. An absolute low‐level light emission standard in the form of the luminol chemiluminescence reaction, either in aqueous or in the non‐aqueous solvent dimethylsulfoxide is proposed for general use and the conditions for its use are described in detail. It should therefore be possible for any researcher to obtain an absolute calibration, in his own experimental geometry, in the blue region of the spectrum.

Melanoma: linked temporal and latitude changes in the United States

The rise in the incidence and mortality from melanoma of the skin is slowing down in younger age groups in the United States. In many White populations, including that of the US, melanoma incidence and mortality rates increase according to proximity of residence to the Equator. Variations with age in this gradient do not seem to have been examined. We examined how the influence of latitude on melanoma rates varied with age. Estimates of age-specific trends by time and by latitude for natural logarithm (Ln) melanoma incidence-rates from the Surveillance, Epidemiology and End Results (SEER) programs, and Ln melanoma mortality rates from the US Vital Statistics were derived from fitted regression equations. Unexpectedly, a decline from old age to youth in the influence of latitude was found for both incidence and mortality from melanoma of the skin in males, and for mortality in females. Further, these changes in the relationship to latitude with age correlated with the changes in time trends with age. The link with exposure suggests that the time trends in melanoma are driven by variations in damage to melanocytes in early life that increases sensitivity to sunlight. This has implications for the general understanding of melanoma etiology and for health education.

Spectral characteristics of the excited states of the product of the chemiluminescence of luminol.

Abstract— In dimethylsulfoxide the emission spectrum of luminol chemiluminescence is red‐shifted by 300 cm‐1 from the photoexcited fluorescence of the product 3‐aminophthalate dianion, while in aqueous solvent the two spectra are identical. The spectral properties of the product dianion have been measured in aqueous solvent and in a number of aprotic solvents, both at room temperature and at 77°K. The ground states and the excited states from which emissions are observed are characterized. Two alternatives are presented to explain the aprotic emission spectra.

THE RELATIONSHIP BETWEEN MALIGNANT MELANOMA OF SKIN AND EXPOSURE TO SUNLIGHT

–Malignant melanomas of the skin are becoming more common. Earlier diagnosis has led to better individual prognoses, but this has not prevented the death rate in the population from rising. This paper brings up‐to‐date studies on the etiology of malignant melanoma, and supplements various fuller but earlier reviews.

Trends in melanoma incidence and mortality

Abstract Many years ago, as part of a general survey of mortality in the United States, 1 it was shown that mortality from all skin cancers combined was rising in the younger age groups. There was apparently an earlier perception in Australia that melanoma was more frequent in the 1930s than it had been in earlier decades, but this is difficult to document. At the time of the U.S. study, the current statistical classification did not separate the cutaneous melanomas from the other skin tumors. Once this separation had been made for a number of years, it was possible to confirm that the rise was due to melanoma. 2

Nevi and migration within the United States and Canada: a population-based cross-sectional study

A survey to ascertain factors associated with benign melanocytic nevi or moles was conducted among randomly-selected White adults (aged 18 to 50 years) in Washington State (United States). Participants of the telephone interview in 1990–91 were questioned about lifetime places of residence and constitutional factors. Subjects counted raised nevi on their arms at the end of the survey. Logistic regression was used to examine the risk for two or more nevi compared with no nevi. Individuals who resided in warmer areas and lower latitudes than Washington State were at higher risk of having multiple nevi. This association held for residence at birth, during childhood, adolescence, and over lifetime: an odds ratio (OR) of 2.3 (95 percent confidence interval =1.2–4.3) for lifetime average daily maximum temperature of ≥64°F compared with 58.9°F, and similar ORs of 2.1 for adolescence and 1.8 for childhood. These associations remained significant after adjusting for potential confounding effects of constitutional factors and for childhood sunburns as a potential mechanism. Risk of multiple nevi was reduced for both early age at migration and longer duration of stay in Washington. These data are consistent with the importance of childhood and adolescent sun exposure in the etiology of nevi, but also suggest an effect of lifetime sun exposure.

The HDAC inhibitor AR42 interacts with pazopanib to kill trametinib/dabrafenib-resistant melanoma cells in vitro and in vivo

Studies focused on the killing of activated B-RAF melanoma cells by the histone deacetylase (HDAC) inhibitor AR42. Compared to other tumor cell lines, PDX melanoma isolates were significantly more sensitive to AR42-induced killing. AR42 and the multi-kinase inhibitor pazopanib interacted to activate: an eIF2α–Beclin1 pathway causing autophagosome formation; an eIF2α–DR4/DR5/CD95 pathway; and an eIF2α-dependent reduction in the expression of c-FLIP-s, MCL-1 and BCL-XL. AR42 did not alter basal chaperone activity but increased the ability of pazopanib to inhibit HSP90, HSP70 and GRP78. AR42 and pazopanib caused HSP90/HSP70 dissociation from RAF-1 and B-RAF that resulted in reduced ‘RAF’ expression. The drug combination activated a DNA-damage-ATM-AMPK pathway that was associated with: NFκB activation; reduced mTOR S2448 and ULK-1 S757 phosphorylation; and increased ULK-1 S317 and ATG13 S318 phosphorylation. Knock down of PERK, eIF2α, Beclin1, ATG5 or AMPKα, or expression of IκB S32A S36A, ca-mTOR or TRX, reduced cell killing. AR42, via lysosomal degradation, reduced the protein expression of HDACs 2/5/6/10/11. In vivo, a 3-day exposure of dabrafenib/trametinib resistant melanoma cells to the AR42 pazopanib combination reduced tumor growth and enhanced survival from ∼25 to ∼40 days. Tumor cells that had adapted through therapy exhibited elevated HGF expression and the c-MET inhibitor crizotinib enhanced AR42 pazopanib lethality in this evolved drug-resistant population.

Neuroimmune mechanisms of behavioral alterations in a syngeneic murine model of human papilloma virus-related head and neck cancer

Patients with cancer often experience a high symptom burden prior to the start of treatment. As disease- and treatment-related neurotoxicities appear to be additive, targeting disease-related symptoms may attenuate overall symptom burden for cancer patients and improve the tolerability of treatment. It has been hypothesized that disease-related symptoms are a consequence of tumor-induced inflammation. We tested this hypothesis using a syngeneic heterotopic murine model of human papilloma virus (HPV)-related head and neck cancer. This model has the advantage of being mildly aggressive and not causing cachexia or weight loss. We previously showed that this tumor leads to increased IL-6, IL-1β, and TNF-α expression in the liver and increased IL-1β expression in the brain. The current study confirmed these features and demonstrated that the tumor itself exhibits high inflammatory cytokine expression (e.g., IL-6, IL-1β, and TNF-α) compared to healthy tissue. While there is a clear relationship between cytokine levels and behavioral deficits in this model, the behavioral changes are surprisingly mild. Therefore, we sought to confirm the relationship between behavior and inflammation by amplifying the effect using a low dose of lipopolysaccharide (LPS, 0.1mg/kg). In tumor-bearing mice LPS induced deficits in nest building, tail suspension, and locomotor activity approximately 24h after LPS. However, these mice did not display an exacerbation of LPS-induced weight loss, anorexia, or anhedonia. Further, while heightened serum IL-6 was observed there was minimal priming of liver or brain cytokine expression. Next we sought to inhibit tumor-induced burrowing deficits by reducing inflammation using minocycline. Minocycline (∼50mg/kg/day in drinking water) was able to attenuate tumor-induced inflammation and burrowing deficits. These data provide evidence in favor of an inflammatory-like mechanism for the behavioral alterations associated with tumor growth in a syngeneic murine model of HPV-related head and neck cancer. However, the inflammatory state and behavioral changes induced by this tumor clearly differ from other forms of inflammation-induced sickness behavior.

A consensus-hemagglutinin-based vaccine delivered by an attenuated Salmonella mutant protects chickens against heterologous H7N1 influenza virus

H7N3 and H7N7 are highly pathogenic avian influenza (HPAI) viruses and have posed a great threat not only for the poultry industry but for the human health as well. H7N9, a low pathogenic avian influenza (LPAI) virus, is also highly pathogenic to humans, and there is a great concern that these H7 subtypes would acquire the ability to spread efficiently between humans, thereby becoming a pandemic threat. A vaccine candidate covering all the three subtypes must, therefore, be an integral part of any pandemic preparedness plan. To address this need, we constructed a consensus hemagglutinin (HA) sequence of H7N3, H7N7, and H7N9 based on the data available in the NCBI in early 2012-2015. This artificial sequence was then optimized for protein expression before being transformed into an attenuated auxotrophic mutant of Salmonella Typhimurium, JOL1863 strain. Immunizing chickens with JOL1863, delivered intramuscularly, nasally or orally, elicited efficient humoral and cell mediated immune responses, independently of the route of vaccination. Our results also showed that JOL1863 deliver efficient maturation signals to chicken monocyte derived dendritic cells (MoDCs) which were characterized by upregulation of costimulatory molecules and higher cytokine induction. Moreover, immunization with JOL1863 in chickens conferred a significant protection against the heterologous LPAI H7N1 virus challenge as indicated by reduced viral sheddings in the cloacal swabs. We conclude that this vaccine, based on a consensus HA, could induce broader spectrum of protection against divergent H7 influenza viruses and thus warrants further study.

Incorporation of membrane-anchored flagellin into Salmonella Gallinarum bacterial ghosts induces early immune responses and protection against fowl typhoid in young layer chickens

The present study aimed to investigate whether the incorporation of flagellin, a TLR5 agonist, in the bacterial ghosts (BGs) of Salmonella Gallinarum can enhance protective immune responses against fowl typhoid, a septicemic disease of poultry, in chickens. BGs are empty cell envelopes derived from Gram-negative bacteria through the bacteriophage phiX174 gene E mediated lysis. In this study, the S. Gallinarum ghosts carrying flagellin were genetically constructed utilizing a lysis plasmid pJHL184-flagellin, designed for the coexpression of the flagellin and the lysis protein E. The adjuvant effect of flagellin was evaluated by immunizing seven day old brown nick layer chicks once orally with either S. Gallinarum-flagellin (SG-fliC) ghosts or S. Gallinarum (SG) ghosts alone. Our results showed that immunization with the SG-fliC ghosts elicited early and higher systemic (IgG) and mucosal (IgA) antibody responses compared to the SG ghosts alone, although not always statistically significant. Flow cytometric analysis of the CD3 + CD4+ and the CD3 + CD8+ T cell populations in peripheral blood mononuclear cells were higher in chickens immunized with the SG-fliC ghosts compared to the chickens vaccinated with the SG ghosts alone. Furthermore, the chickens immunized with SG-fliC ghosts exhibited significantly (p < 0.05) higher IL-6 and IFN-γ responses compared to the chickens vaccinated with the SG ghosts alone. On challenge with the virulent S. Gallinarum wild type strain at 28th day post immunization, 5 of 10 birds died (50%) in case of SG-fliC ghost group while 60% (6 of 10 birds died) mortality was observed in the SG ghost group. Collectively, these results suggest that the expression of flagellin in SG ghosts improves antigen-specific humoral and cell mediated immune responses, and can enhance protective efficacy of the BG-based vaccines against the virulent challenges.