In situ biodistribution and residency of a topical anti-inflammatory using fluorescence lifetime imaging microscopy.

BACKGROUND: GSK2894512 is a topically delivered investigational drug being developed for treatment of atopic dermatitis and psoriasis. OBJECTIVES: To investigate, in a phase I clinical trial, the spatial biodistribution and residency of GSK2894512 within the epidermis and dermis of healthy human participants noninvasively using fluorescence lifetime imaging microscopy (FLIM). METHODS: Two topical drug formulations containing GSK2894512 1% were applied to the right and left forearms of six participants for seven consecutive days, followed by seven days of observation for residency. FLIM images were obtained daily throughout the study, approximately every 24 h. During the treatment phase of the study, images were collected from each participant pretreatment, reflecting the residual dose from the previous day. Three punch biopsies from each participant of one formulation was obtained from the treated region during the post-treatment follow-up period between days 8 and 14 for comparison with FLIM results. RESULTS: Cellular and subcellular features associated with different epidermal and dermal layers were visualized noninvasively, down to a depth of 200 µm. Results yielded three-dimensional maps of GSK2894512 spatial distribution and residency over time. This fluorescence data provided a marker that was used as a monitor for day-to-day variance of drug presence and residency postapplication. CONCLUSIONS: The results suggest FLIM could be a viable alternative to skin biopsies without the usual patient discomfort and limitations, thereby enabling the direct measurement of skin distribution through longitudinal monitoring. These results are the first step in establishing the unique capabilities that multiphoton imaging could provide to patients through noninvasive drug detection.

The effect of pH on the dynamics of natural membranes.

Pure phospholipids and membrane fragments from bacterial cells living under various conditions were studied against the influence of the surrounding acidity on the internal dynamics. For that we compared mean square displacements extracted from elastic incoherent neutron scattering data, measured both at low and at neutral pH, of the phospholipids 1,2-dimyristoyl-sn-glycero-3-phosphocholine and of samples from neutralophilic and acidophilic micro-organisms (some being hyperthermophilic and others mesophilic). The lipids showed a slight shift in the phase transition temperature of about 4 degrees under pH variation and became slightly more mobile at lower pH. The membrane fragments not used to extreme acidic conditions were significantly more sensitive to variations in the pH values, whereas the acidophilic and -tolerant samples were much less influenced by this parameter. They presented the higher softness at low pH, which was closer to their native condition. Such findings might be a hint for adaptation mechanisms to different acidity conditions.

A single exposure to hyperbaric oxygen increases levels of circulating nucleosomes but does not induce mononuclear cell apoptosis in divers.

Recent reports that hyperbaric oxygenation (HBO2) induced apoptosis in T-cell lines raised concern about a possible immunosuppressive effect of HBO2. Nucleosomes, DNA fragments wrapped around a histone core, have been observed in the circulation in diseases with increased cell death such as sepsis. Our aim was to investigate, whether HBO2 increases circulating nucleosomes as a marker of cell death and induces apoptosis of peripheral blood mononuclear cells in vivo. After informed consent 29 healthy volunteers were exposed to a 30 minute dive at 2.8 atmospheres absolute in a pressure chamber under resting conditions, while breathing 100% oxygen. Samples were obtained before and 24 hours after exposure. Circulating nucleosomes were measured in serum. Caspase-3 activation, Bcl-2 expression and mRNA of Bcl-2, Bcl-xl and Bax were analyzed in mononuclear cell extracts. Nucleosomes were elevated markedly 24h after exposure (p<0.01), while caspase-3 was not activated significantly. mRNA levels of Bcl-2, Bcl-xl and Bax were not altered. In conclusion, while evidence of elevated levels of circulating nucleosomes was found, mononuclear cell apoptosis was not affected by a single exposure to hyperbaric oxygen.

Angiogenesis correlates with metastasis in melanoma.

BACKGROUND: Angiogenesis has been correlated with melanoma progression, but its role in melanoma metastasis is unclear. METHODS: To determine whether angiogenesis correlates with the presence of melanoma metastases, we compared the number of microvessels in the primary melanomas of 12 patients presenting with metastases to those of 13 patients without metastases. Patient groups were matched for gender, age, tumor depth, and histological type and anatomical location of the primary melanoma. Microvessels were stained with factor VIII antibody and counted. RESULTS: Microvessel counts were significantly greater for the metastatic than the nonmetastatic melanomas (51.63+/-14.95 vs. 24.86+/-8.415; P < .0001). One hundred percent of the metastatic melanomas had a mean microvessel count of > or = 37, whereas only 8% of the nonmetastatic melanomas had a mean microvessel count of > or = 37 (sensitivity = 1.00, specificity = .92). Interestingly, patients with lymph node metastases had significantly lower microvessel counts than did patients with distant metastases (42.00+/-3.482 vs. 58.50+/-16.40; P < .05), and significantly higher microvessel counts than did patients without metastases (42.00+/-3.482 vs. 24.86+/-8.415; P < .001). CONCLUSIONS: An increased number of microvessels in the primary tumors of patients with melanoma correlates with the simultaneous presence of metastases. This suggests that angiogenesis may be important in the process of melanoma metastasis.