Label-Free Quantification (LFQ) of Fecal Proteins for Potential Pregnancy Detection in Polar Bears.

Reliable pregnancy diagnostics would be beneficial for monitoring polar bear (Ursus maritimus) populations both in situ and ex situ, but currently there is no method of non-invasive pregnancy detection in this species. Recent reports in several carnivore species described the identification of fecal proteins that may serve as pregnancy biomarkers; however, repeatability has been limited. The objective of the current analysis was to utilize an unbiased, antibody-free, label-free method for the identification and quantification of fecal proteins to determine if differences associated with pregnancy are detectable in polar bears. Protein was extracted from fecal samples (n = 48) obtained from parturient (n = 6) and non-parturient (n = 6) profiles each at four timepoints: pre-breeding season, embryonic diapause, early placental pregnancy, and mid-placental pregnancy. Protein was prepared and analyzed on the Thermo Orbitrap Eclipse nanoLC-MS/MS system. A total of 312 proteins was identified and quantified; however, coefficients of variation (CV) were high for both abundance ratio variability (384.8 ± 61.0% SEM) and within group variability (86.8 ± 1.5%). Results of this study suggest that the inconsistencies in specific protein concentrations revealed previously by antibody-based assays may not be due to that methodology's limitations, but rather, are reflective of true variation that exists among samples.

Cutting Edge: l-Arginine Transfer from Antigen-Presenting Cells Sustains CD4+ T Cell Viability and Proliferation.

Metabolomics analyses suggest changes in amino acid abundance, particularly l-arginine (L-ARG), occur in patients with tuberculosis. Immune cells require L-ARG to fuel effector functions following infection. We have previously described an L-ARG synthesis pathway in immune cells; however, its role in APCs has yet to be uncovered. Using a coculture system with mycobacterial-specific CD4+ T cells, we show APC L-ARG synthesis supported T cell viability and proliferation, and activated T cells contained APC-derived L-ARG. We hypothesize that APCs supply L-ARG to support T cell activation under nutrient-limiting conditions. This work expands the current model of APC-T cell interactions and provides insight into the effects of nutrient availability in immune cells.

Biotherapy of Brain Tumors with Phosphatidylserine-Targeted Radioiodinated SapC-DOPS Nanovesicles.

Glioblastoma multiforme (GBM), a common type of brain cancer, has a very poor prognosis. In general, viable GBM cells exhibit elevated phosphatidylserine (PS) on their membrane surface compared to healthy cells. We have developed a drug, saposin C-dioleoylphosphatidylserine (SapC-DOPS), that selectively targets cancer cells by honing in on this surface PS. To examine whether SapC-DOPS, a stable, blood-brain barrier-penetrable nanovesicle, could be an effective delivery system for precise targeted therapy of radiation, we iodinated several carbocyanine-based fluorescent reporters with either stable iodine (127I) or radioactive isotopes (125I and 131I). While all of the compounds, when incorporated into the SapC-DOPS delivery system, were taken up by human GBM cell lines, we chose the two that best accumulated in the cells (DiI (22,3) and DiD (16,16)). Pharmacokinetics were conducted with 125I-labeled compounds and indicated that DiI (22,3)-SapC-DOPS had a time to peak in the blood of 0.66 h and an elimination half-life of 8.4 h. These values were 4 h and 11.5 h, respectively, for DiD (16,16)-SapC-DOPS. Adult nude mice with GBM cells implanted in their brains were treated with 131I-DID (16,16)-SapC-DOPS. Mice receiving the radionuclide survived nearly 50% longer than the control groups. These data suggest a potential novel, personalized treatment for a devastating brain disease.

Clearance in vivo of instilled [h]cholesterol from the rat lung.

Phospholipids and lung surfactant proteins are known to be recycled within the lung alveolus mainly by uptake into type II epithelial cells that secrete lipid-enriched lung surfactant. Dipalmitoyl phosphatidylcholine (DPPC) is the major component of lung surfactant lipids and cholesterol is the second most abundant. However, cholesterol turnover in vivo has not been measured and it is not known how long steroidal compounds persist in the lung in intact animals. Here we report on experiments in which radiolabeled cholesterol was instilled into the lungs of rats, then at various postinstillation periods, radioactive sterols in lavage fluid, and in postlavage whole lungs were measured in individual animals. Radioactive sterols in the lungs remained high for a week and were still detectable 46 days later. The clearance rate during the initial postinstillation week was approximately 10% per day. Both radioactive free and esterified sterols were recovered from bronchoalveolar lavage fluid and postlavage lungs.

Evidence for DNA synthesis in Pneumocystis carinii trophozoites treated with the beta-1,3-glucan synthesis inhibitor pneumocandin L-693,989.

Pneumocandins inhibit beta-1,3-glucan synthesis preventing the development of Pneumocystis cysts that are absent from the lungs of treated rats. To determine whether treated trophozoites are capable of DNA replication, cytochemical analyses were performed on 4',6-diamidino-2-phenylindole (DAPI)- and DB181-stained Pneumocystis carinii isolated from pneumocandin L-693-989-treated rats. Fluorescence intensities of trophozoite nuclei from drug-treated rats were greater than those of untreated controls, suggesting that DNA replication was not inhibited but that cytokinesis and perhaps karyokinesis were blocked.