Desulfurization of model and diesel oils by resting cells of Gordona sp.

The desulfurization activity of the resting cells of Gordona sp. CYKS1 was strongly depended on harvest time and the highest value when the cells had been harvested in the early growth phase (0.12 mg sulfur g^–1 cell^–1 h^–1). For the model oil, hexadecane containing dibenzothiophene, the specific desulfurization rate decreased as the reaction proceeded. Both the specific and the volumetric desulfurization rates were not significantly affected by the aqueous-to-oil phase ratio. The diesel oils, light gas oil and a middle distillate unit feed were desulfurized at higher rates (ca. 0.34 mg sulfur g^–1 cell^–1 h^–1) than the model oil (0.12 mg sulfur g^–1 cell^–1 h^–1).     

Enhancement of cell growth by intracellularly expressed herpes simplex virus thymidine kinase

A recombinant cell line (NIH3T3:pLtkSN) was made by infecting parental cells (NIH3T3) with a recombinant retrovirus (pLtkSN) encoding herpes simplex virus thymidine kinase (HSVtk) gene. The cells expressing HSVtk (NIH3T3:pLtkSN) grew 2.3 times more than the parental cells (NIH3T3) in Dulbecco's Modified Eagles Media containing 10% (v/v) horse serum. The NIH3T3:pLtkSN cells also showed a significant enhancement in the maximal cell concentration and the specific growth rate even at 2.5% serum concentration. The specific O2 uptake rate of NIH3T3 was 2.1 times greater than that of NIH3T3:pLtkSN. Under both O2-limited and O2-unlimited conditions, it appears that HSVtk plays an important role in enhancing the growth characteristics of animal cells.     

Regulation of G(1) cell-cycle progression by oncogenes and tumor suppressor genes.

Progression of resting quiescent G(0) cells into early G(1) and transition across the restriction point are highly regulated processes. Mutation of proto-oncogenes and tumor suppressor genes regulating these transitions are targeted during oncogenesis. Recent work has underscored the importance of the G(0) to early G(1) transition and metabolism to neoplastic cells.     

Human beta-glucuronidase. Studies on the effects of pH and bile acids in regard to its role in the pathogenesis of cholelithiasis

Human bile contains a considerable amount of endogenous beta-glucuronidase. The effects of pH and bile acids on its activity have been studied in regard to its role in the pathogenesis of cholelithiasis. beta-Glucuronidase, purified from human liver to homogeneity, was structurally stable between pH 4 and 10, but was active only over a much narrower range of pH, with a pH optimum of 5.2. The inactivation below pH 4 was due to its irreversible denaturation, whereas the inactivation at higher pH was due to a true reversible pH effect on the enzyme velocity. Kinetic studies revealed that hydrogen ion acted as a substrate-directed activator of the free enzyme, but not the enzyme-substrate complex, with a molecular dissociation constant of 4 X 10(-6). The enzyme activity was not affected by unconjugated bile acids, primarily due to their extremely low water solubility. Conjugated bile acids, on the other hand, exerted heterogeneous and pH-dependent effects on the enzyme. At pH 5.2, taurocholic acid and glycocholic acid were substrate-directed activators of the enzyme; taurochenodeoxycholic acid and taurodeoxycholic acid, competitive inhibitors; and glycochenodeoxycholic acid and glycodeoxycholic acid, mixed inhibitors. At pH 7.0 all taurine and glycine conjugates behaved as substrate-directed activators. Though beta-glucuronidase activity at pH 7 was only 23% of its maximal activity at pH 5.2, conjugated bile acids tended to restore its activity to a certain extent at pH 7. Thus, endogenous beta-glucuronidase could play a significant role in pigment cholelithiasis.     

Relationship between postabsorptive respiratory exchange ratio and plasma free fatty acid concentrations

The relationship between overnight postabsorptive (fasting) respiratory exchange ratio (RER) and plasma FFA concentrations was addressed using data from three separate protocols, each of which involved careful control of the antecedent diet. Protocol 1 examined the relationship between fasting RER and the previous daytime RER. In Protocol 2 fasting, RER and plasma palmitate concentrations were measured in 29 women and 31 men (body mass index <30 kg·m⁻²). Protocol 3 analyzed data from Nielsen et al. (Nielsen, S., Z. K. Guo, J. B. Albu, S. Klein, P. C. O''Brien, M. D. Jensen. 2003. Energy expenditure, sex and endogenous fuel availability in humans. J. Clin. Invest. 111: 981-988.) to understand how fasting RER and palmitate concentrations relate within individuals during four consecutive measurements. The results were as follows: 1) Fasting RER was correlated (r = 0.74, P < 0.001) with the previous day''s average RER, and less so with RER variability. 2) Fasting RER was correlated (r = −0.39, P = 0.007) with fasting plasma palmitate concentrations. 3) The pattern of the RER/palmitate relationship was similar within individuals and between individuals; a negative slope was observed significantly more often than a positive slope (χ² test; P < 0.001). Our findings suggest that, despite a fixed food quotient, the slight departures from energy equilibrium in a controlled General Clinical Research Center environment can effect plasma FFA concentrations. We suggest that including indirect calorimetry as part of FFA metabolism studies may aid in data interpretation.     

Primate stem cells: bridge the translation from basic research to clinic application

A growing body of literature has shown that stem cells are very effective for the treatment of degenerative diseases in rodents but these exciting results have not translated to clinical practice. The difference results from the divergence in genetic, metabolic, and physiological phenotypes between rodents and humans. The high degree of similarity between non-human primates(NHPs) and humans provides the most accurate models for preclinical studies of stem cell therapy. Using a NHP model to understand the following key issues, which cannot be addressed in humans or rodents, will be helpful for extending stem cell applications in the basic science and the clinic. These issues include pluripotency of primate stem cells, the safety and efficiency of stem cell therapy, and transplantation procedures of stem cells suitable for clinical translation. Here we review studies of the above issues in NHPs and current challenges of stem cell applications in both basic science and clinical therapies. We propose that the use of NHP models, in particular combining the serial production and transplantation procedures of stem cells is the most useful for preclinical studies designed to overcome these challenges.     

Composition and peptide maps of cross-linked human choriogonadotropin-receptor complexes on porcine granulosa cells

Radioiodinated human choriogonadotropin was affinity-cross-linked with a cleavable (nondisulfide) homobifunctional reagent to the hormone receptor on porcine granulosa cells and the solubilized sample was electrophoresed. Cross-linked samples revealed four additional bands of slower electrophoretic mobility in addition to the hormone alpha, beta, and alpha beta dimer bands. The four bands corresponded to masses of 68, 74, 102, and 136 kDa whereas the alpha beta dimer band corresponded to 50 kDa. Formation of the four bands requires the 125I-hormone to bind specifically to the receptor with subsequent cross-linking. Binding can be prevented by excess of native hormone but not by follitropin. A monofunctional analog of the cross-linking reagent failed to produce the four bands. They were also produced by cross-linking Triton X-100-solubilized hormone-receptor complexes. Reagent concentration-dependent cross-linking revealed that their formation was sequential; smaller complexes formed first and then larger ones. When gels of the cross-linked sample were treated with reagents that cleave covalent cross-links and then electrophoresed in a second dimension gel, 18-, 24-, 28-, and 34-kDa components were released, in addition to the alpha and beta subunits of the native hormone. Simultaneous peptide mapping of the cross-linked complexes in the gel matrix with Staphylococcus V8 protease or papain revealed progressive proteolysis to generate terminal fragments of 30 or 27 kDa, respectively. These fragments were unique to and commonly present in the 74-, 102-, and 136-kDa hormone-receptor complexes but were not produced by proteolysis of the cross-linked human choriogonadotropin (hCG) alpha beta dimer or the hCG alpha subunit. Apparently, the radioactively labeled segment(s) of the alpha subunit of 125I-hCG was cross-linked to the 24-kDa component. The results demonstrate the protein nature of the receptor and suggest that 125I-hCG was initially cross-linked to the 24-kDa component to generate the 74-kDa complex, then the 28- and 34-kDa components were sequentially cross-linked to the 24-kDa component in the 74-kDa complex to generate the 102- and 134-kDa complexes.     

Sex differences in the electrocommunication signals of the electric fish Apteronotus bonapartii

The South American weakly-electric knifefish (Apteronotidae) produce highly diverse and readily quantifiable electrocommunication signals. The electric organ discharge frequency (EODf), and EOD modulations (chirps and gradual frequency rises (GFRs)), vary dramatically across sexes and species, presenting an ideal opportunity to examine the proximate and ultimate bases of sexually dimorphic behavior. We complemented previous studies on the sexual dimorphism of apteronotid communication signals by investigating electric signal features and their hormonal correlates in Apteronotus bonapartii, a species which exhibits strong sexual dimorphism in snout morphology. Electrocommunication signals were evoked and recorded using a playback paradigm, and were analyzed for signal features including EOD frequency and the structure of EOD modulations. To investigate the androgenic correlates of sexually dimorphic EOD signals, we measured plasma concentrations of testosterone and 11-ketotestosterone. A. bonapartii responded robustly to stimulus playbacks. EODf was sexually monomorphic, and males and females produced chirps with similar durations and amounts of frequency modulation. However, males were more likely than females to produce chirps with multiple frequency peaks. Sexual dimorphism in apteronotid electrocommunication signals appears to be highly evolutionarily labile. Extensive interspecific variation in the magnitude and direction of sex differences in EODf and in different aspects of chirp structure suggest that chirp signals may be an important locus of evolutionary change within the clade. The weakly-electric fish represent a rich source of data for understanding the selective pressures that shape, and the neuroendocrine mechanisms that underlie, diversity in the sexual dimorphism of behavior.     

Human P301L-Mutant Tau Expression in Mouse Entorhinal-Hippocampal Network Causes Tau Aggregation and Presynaptic Pathology but No Cognitive Deficits

Accumulation of hyperphosphorylated tau in the entorhinal cortex (EC) is one of the earliest pathological hallmarks in patients with Alzheimer’s disease (AD). It can occur before significant Aβ deposition and appears to “spread” into anatomically connected brain regions. To determine whether this early-stage pathology is sufficient to cause disease progression and cognitive decline in experimental models, we overexpressed mutant human tau (hTauP301L) predominantly in layer II/III neurons of the mouse EC. Cognitive functions remained normal in mice at 4, 8, 12 and 16 months of age, despite early and extensive tau accumulation in the EC. Perforant path (PP) axon terminals within the dentate gyrus (DG) contained abnormal conformations of tau even in young EC-hTau mice, and phosphorylated tau increased with age in both the EC and PP. In old mice, ultrastructural alterations in presynaptic terminals were observed at PP-to-granule cell synapses. Phosphorylated tau was more abundant in presynaptic than postsynaptic elements. Human and pathological tau was also detected within hippocampal neurons of this mouse model. Thus, hTauP301L accumulation predominantly in the EC and related presynaptic pathology in hippocampal circuits was not sufficient to cause robust cognitive deficits within the age range analyzed here.