HARBOR PORPOISE (PHOCOENA PHOCOENA) ABUNDANCE IN ALASKA: BRISTOL BAY TO SOUTHEAST ALASKA, 1991-1993

Between 1991 and 1993, Alaska harbor porpoise ( Phocoena phocoena ) abundance was investigated during aerial surveys throughout much of the coastal and offshore waters from Bristol Bay in the eastern Bering Sea to Dixon Entrance in Southeast Alaska. Line-transect methodology was used, and only those observations made during optimal conditions were analyzed. Survey data indicated densities of 4.48 groups/100 km², or approximately 3,531 harbor porpoises (95% C. I. 2,206-5,651) in Bristol Bay and 0.54 groups/100 km², or 136 harbor porpoises (95% C. I. 11-1,645) for Cook Inlet. Efforts off Kodiak Island resulted in densities of 1.85 groups/100 km², or an abundance estimate of 740 (95% C. I. 259-2,115). Surveys off the south side of the Alaska Peninsula found densities of 2.03 groups/100 km² and an abundance estimate of 551 (95% C. I. 423-719). Surveys of offshore waters from Prince William Sound to Dixon Entrance yielded densities of 4.02 groups/100 km² and an abundance estimate of 3,982 (95% C. I. 2,567-6,177). Combining all years and areas yielded an uncorrected density estimate of 3.82 porpoises per 100 km², resulting in an abundance estimate of 8,940 porpoises (CV = 13.8%) with a 95% confidence interval of 6,746-11,848. Using correction factors from other studies to adjust for animals missed by observers, the total number of Alaska harbor porpoises is probably three times this number.     

Extended type 1 chain glycosphingolipids: dimeric Lea (III4V4Fuc2Lc6) as human tumor-associated antigen

Glycolipid extracts from various human cancer tissues and cell lines showed the presence of a slow-migrating glycolipid component which was strongly reactive with monoclonal antibody (mAb) NCC-ST-421 (raised against human gastric adenocarcinoma) and weakly cross-reactive with anti-Lea mAbs. The slow-migrating glycolipid was isolated from human colonic adenocarcinoma cell line Colo205 grown in nude mice, and was purified by high-performance liquid chromatography followed by preparative thin-layer chromatography. Its structure was elucidated by sequential enzymatic degradation and thin-layer chromatography immunostaining of the degradation products with various mAbs, 1H NMR spectroscopy, positive-ion fast atom bombardment mass spectrometry, and methylation analysis. The major slow-migrating component reacting with mAb ST-421 was identified as dimeric Lea, with the structure as follows. [formula: see text] Antigens containing this structure and various analogous structures (including enzymatically synthesized Lea/Lex hybrid antigen) were tested with ST-421. While the mAb was equally reactive with dimeric Lea and Lea/Lex, only the former was chemically detectable as the slow-migrating glycolipid from the tumor extract. ST-421 showed less reactivity with simple Lea (III4FucLc4) or extended Lea (V4FucLc6, and/or IV3Gal beta 1----3[Fuc alpha 1----4]GlcNAcnLc4), and was not reactive with Lex/Lex (dimeric Lex). It was concluded, therefore, that the major tumor-associated slow-migrating glycolipid reacting with ST-421 has the dimeric Lea structure shown above. Since extension of lacto-series structure has been shown to be limited to type 2 chain in normal cells and tissues, extended elongation of type 1 chain as shown in this structure represents a novel tumor-associated epitope.     

Intralesional lipid-complexed cytokine/superantigen immunogene therapy for spontaneous canine tumors

These studies sought to determine the gene expression and short-term effects of intralesional lipid-complexed immunogene therapy with constructs encoding Staphylococcus aureus enterotoxin A and canine interleukin-2 (L-SEA/cIL-2) in dogs with tumors of various histotypes, and then to assess the safety and efficacy of repeated L-SEA/cIL-2 injections in dogs with spontaneous soft tissue sarcomas (STS). In the first study, pet dogs with a variety of tumors received a single intralesional injection of L-SEA/cIL-2, and surgical excision was performed 48 h later. In the second study, dogs with histologically confirmed STS were treated weekly for a maximum of 12 weeks with escalating doses of L-SEA/cIL-2. Tumors were then surgically excised and assessed histologically and immunohistochemically. Overall, treatments were well tolerated, with no dose-limiting toxicities encountered. At 48 h, in the single injection study, plasmid DNA was detected in 14 of 16 tumor samples, and plasmid-specific mRNA was detected in 3 of 14. In the multiple injection study, the overall response rate in dogs with STS was 25%, consisting of 3 complete responses (CR) and 1 partial response (PR). Diffuse lymphoplasmacytic inflammation was observed in all tumors from patients experiencing CR or PR, whereas these changes were not evident in tumors from nonresponders. The infiltrate was composed primarily of CD3(+) cells at 48 h from the single-injection study, and was composed of both CD3(+) and CD79a(+) cells at 12 weeks in responding dogs from the multiple-injection study. In conclusion, these studies suggests that intralesional L-SEA/cIL-2 immunotherapy is well tolerated, results in detectable transgene expression in canine tumors, and has antitumor activity in dogs with spontaneous STS.     

Androgen metabolism in tissue recombinants composed of adult urinary bladder epithelium and urogenital sinus mesenchyme

Epithelium of the adult mouse urinary bladder (BLE) was experimentally combined with mesenchyme of the urogenital sinus (UGM) and grown in intact male hosts to produce prostate-like glandular structures. To determine the extent to which the BLE is altered in a functional sense by inductive influences from UGM, investigations into the in vitro metabolism of tritiated testosterone (T) were undertaken. An isocratic high performance liquid chromatographic (HPLC) method was developed in order to separate the metabolites of T in mouse bladder, prostate and UGM + BLE tissue recombinants. Using a C-18 reversed phase column and a tetrahydrofuran (20): methanol (40): H2O (40) mobile phase, efficient and rapid separation of T, dihydrotestosterone, 3 alpha-androstanediol, androstenedione, androstanedione and androsterone was achieved. The identities of the radiolabeled T metabolites were confirmed by recrystallization to constant specific activity. The results of the present study revealed that tissue recombinants expressed testosterone metabolic profiles only partially toward that of the adult prostate. For example, percentage formation of 5 alpha-androstanedione, 3 alpha-androstanediol and unknown polar metabolites in the UGM + BLE resembled the prostate and differed significantly from the urinary bladder. Conversely, formation of the 3 beta-androstanediol and androsterone from testosterone resembled the urinary bladder and differed from the formation of these metabolites in the prostate. These results suggest that in contrast to histomorphology, androgen-induced DNA synthesis, androgen receptor binding activity and total tissue two-dimensional gel electrophoretic protein profiles, androgen metabolic profiles in the tissue recombinants showed only partial transformation into prostatic phenotypes. Analysis of steroid-metabolic profiles, therefore, may represent an exquisite and sensitive method to assess gene expression in various hormone-responsive target tissues.     

Maternal, fetal, and newborn tissue PO2 in sheep measured with galvanic oxygen electrodes

Tissue PO2 was measured with galvanic oxygen electrodes chronically implanted in maternal and fetal tissues of two pregnant ewes. Labor commenced after 3 days in one ewe and after 11 days in the other. Tissue PO2 was measured in both newborn lambs. Fetal tissue PO2 during labor was less than 3 mm Hg but rose to values similar to those found in adult tissues in the newborn period. Function of the electrode and response of the tissue to change in oxygenation was challenged by administration of 100% oxygen to the pregnant ewe or newborn lamb, and by administration of intravenous epinephrine to the ewe. These tests suggested that all electrodes were functioning satisfactorily in vivo and that tissue PO2 was responding as anticipated. Recalibration of electrodes after removal from the tissues confirmed that their response in vitro was similar to that obtained before implantation.     

Detection of radical adducts of 5,5-dimethyl-1-pyrroline n-oxide by the combined use of high-performance liquid chromatography with electrochemical detection and electron spin resonance.

Although high-performance liquid chromatography with electrochemical detection (HPLC-EC) has been previously used to analyze radicals trapped by 5,5-dimethyl-1-pyrroline-N-oxide (DMPO), these techniques omitted the critical use of an internal standard. To improve reliability we have incorporated 3-carbamoyl-2,2,5,5-tetramethylpyrrolidin-1-yloxy free radical (CTPO) as a stable, oxidizable, and paramagnetic internal standard. Radicals generated during the uv photolysis of hydrogen peroxide were trapped by DMPO, analyzed by HPLC-EC, and quantified by relative comparison of peak areas to those of CTPO. DMPO adducts of hydroxyethyl, hydroxymethyl, and carbon dioxide anion radicals have been further characterized by ESR analysis of their 13C isotopes sampled from the HPLC eluant. Studies of the voltage dependence of the electrochemical signal demonstrate how this can be used to further confirm the identity of artifactual HPLC-EC peaks with similar retention times.