Fructose bisphosphatase from Escherichia coli. Purification and characterization

Escherichia coli fructose-1,6-bisphosphatase has been purified for the first time, using a clone containing an approximately 50-fold increased amount of the enzyme. The procedure includes chromatography in phosphocellulose followed by substrate elution and gel filtration. The enzyme has a subunit molecular weight of approximately 40,000 and in nondenaturing conditions is present in several aggregated forms in which the tetramer seems to predominate at low enzyme concentrations. Fructose bisphosphatase activity is specific for fructose 1,6-bisphosphate (Km of approximately 5 microM), shows inhibition by substrate above 0.05 mM, requires Mg2+ for catalysis, and has a maximum of activity around pH 7.5. The enzyme is susceptible to strong inhibition by AMP (50% inhibition around 15 microM). Phosphoenolpyruvate is a moderate inhibitor but was able to block the inhibition by AMP and may play an important role in the regulation of fructose bisphosphatase activity in vivo. Fructose 2,6-bisphosphate did not affect the rate of reaction.     

An IS4 transposition causes a 13-bp duplication of phage lambda DNA and results in the constitutive expression of the cI and cro gene products

We have examined the nature of the additional DNA present in lambda hyp- mutants (Eisen et al., 1982). This DNA is an IS4 element in orientation I, in the y region of bacteriophage lambda at nucleotide position 39,139 (see Moore et al., 1979). Our assignment is based on (i) the similarity in size derived from the PstI, AvaI, and HindII restriction pattern and (ii) the DNA sequence of both the left and right lambda-IS4 DNA junctions in phage lambda hyp15rev4. The IS4 integration event resulted in the duplication of 13 bp of lambda DNA in contrast to the 11- and 12-bp duplications previously observed at the sites of IS4 integrations elsewhere (Klaer et al., 1981).     

Parkinson’s disease: clinical aspects

Parkinsonism is a clinical syndrome characterized by akinesia, muscular rigidity, and resting tremor. The most frequent cause of parkinsonism is Parkinsons disease (PD). Progressive loss of substantia nigra neurons together with the occurrence of Lewy bodies are considered essential neuropathological features of PD. Recent neuropathological studies suggest that nigral degeneration is only part of a more extended brain degeneration that starts in the medulla oblongata and then spreads to the mesencephalon and cerebral cortex. Correspondingly, the clinical symptoms occurring in PD go far beyond parkinsonism. Depending on the disease stage, autonomic dysfunction, olfactory disturbances, depression, and dementia are frequently encountered in PD. These neuropathological and clinical observations have major implications for future research in PD. In particular, the analysis of the properties that the neuronal cell types involved in PD have in common and that might make them susceptible to degeneration is essential.     

Ribavirin enhances interferon-γ levels in patients with chronic hepatitis C treated with interferon-α

Some patients with chronic hepatitis C respond to interferon (IFN)-alpha treatment, and the efficiency can be improved by combining it with ribavirin. The mechanism of this improvement is unknown. To investigate the effects of these two regimens on the immune responses in 51 patients with chronic hepatitis C, we examined the hepatitis C core antigen-specific proliferative response and cytokine production profiles, natural killer (NK) cell cytotoxicity and cytotoxic T cell function during treatment. The results are as follows: (1) both viral clearance and biochemical normalization occurred more frequently in patients receiving combination therapy; (2) the function of NK cells increased after treatment in the responders of both groups (p < 0.05); (3) the level of IFN-gamma produced by hepatitis C core antigen-stimulated peripheral blood mononuclear cells was higher in patients receiving combination therapy, especially in responders; (4) the core antigen-specific proliferative response decreased after treatment, and (5) in addition, the core-specific cytotoxic T cell activities of five responder patients also increased significantly after therapy. In conclusion, enhancement of immune responses, especially those related to type-1 T helper cell activity, may contribute to better efficacy in combining ribavirin with IFN-alpha for treatment of chronic hepatitis C.     

藏北高寒草地植被的碳密度与碳贮量

采用实地调查与查阅文献相结合的方法估算藏北高寒草地植被碳密度和碳贮量。结果表明:1)藏北高寒草地总面积约39.059×106 hm²,植被地上平均碳密度12.158±4.7g·m^-2,植被地下平均碳密度84.458±20.38g·m^-2,植被地上部碳贮量5.171±0.95Tg,植被地下部碳贮量25.223±2.96Tg,植被总碳贮量为30.394±3.91Tg;2)不同草地组间植被碳密度和碳贮量差异显著。其中不同草地组间植被碳密度以丛生禾草组碳密度值最低,地上和地下碳密度分别为6.13±1.51g·m^-2和26.04±5.8g·m^-2,具灌木的半灌木组碳密度最高,地上和地下碳密度分别为31±3.4 g·m^-2和244.59±6.9g·m^-2;而不同草地组间,草地植被碳贮量以小莎草组最大,植被地上和地下碳贮量分别为2.24±0.32Tg和9.52±0.89Tg,半灌木组碳贮量最小,其地上和地下碳贮量分别为0.012 4±0.002Tg和0.098 1±0.002Tg。3)藏北高寒草地分布各县(区)碳密度和碳贮量的分布也存在显著差异。从碳密度来看,革吉县、札达县、噶尔县和措勤县碳密度较高,植被平均碳密度分别相当于藏北平均植被碳密度的1.76,1.47,1.11和1.06倍,从碳贮量来看,碳贮量集中分布于双湖特别区、札达、尼玛、日土、革吉和改则6县(区),六县(区)草地植被碳贮量为25.2±2.31Tg,占藏北总植被碳量的82.89%。     

Isolation of oligodendroglial cells from young and adult whole rat brains using an in situ generated percoll density gradient

A method for the isolation of oligodendroglial cells from young and adult whole rat brains, using a Percoll density gradient is presented. The minced tissue, incubated in a balanced salt solution containing 0.1% trypsin is further dissociated by forcing it through nylon screens to 145 and 74 μm pore size. The crude suspension is then mixed with an isosmotic Percoll solution and centrifuged for 15 min. An in situ generated density gradient allows the separation of five bands, only one of which (Band C) lying between δ1.050 and δ1.062 contains cellular elements. The isolated cells show the typical morphological characteristics of oligodendroglia.A detailed morphological study of the cells isolated from whole brains of 10-, 30- and 120-day old rats is presented for the first time in the literature and immunocytochemical characterization is carried out using specific (antigalactocerebroside) and non specific (anti-glial fibrillary acidic protein) anti-sera.The method is simple and rapid and isosmotic conditions are maintained throughout, resulting in a better preservation of cell integrity. It represents an improvement over the two previous methods described in the literature and will be useful for studying different developmental events (biochemical and morphological) occurring in oligodendroglial cells at early stages of myelin formation.     

Activation of Neuropeptide Y Receptors Modulates Retinal Ganglion Cell Physiology and Exerts Neuroprotective Actions In Vitro

Neuropeptide Y (NPY) is expressed in mammalian retina but the location and potential modulatory effects of NPY receptor activation remain largely unknown. Retinal ganglion cell (RGC) death is a hallmark of several retinal degenerative diseases, particularly glaucoma. Using purified RGCs and ex vivo rat retinal preparations, we have measured RGC intracellular free calcium concentration ([Ca²⁺]_i) and RGC spiking activity, respectively. We found that NPY attenuated the increase in the [Ca²⁺]_i triggered by glutamate mainly via Y₁ receptor activation. Moreover, (Leu³¹, Pro³⁴)−NPY, a Y₁/Y₅ receptor agonist, increased the initial burst response of OFF-type RGCs, although no effect was observed on RGC spontaneous spiking activity. The Y₁ receptor activation was also able to directly modulate RGC responses by attenuating the NMDA-induced increase in RGC spiking activity. These results suggest that Y₁ receptor activation, at the level of inner or outer plexiform layers, leads to modulation of RGC receptive field properties. Using in vitro cultures of rat retinal explants exposed to NMDA, we found that NPY pretreatment prevented NMDA-induced cell death. However, in an animal model of retinal ischemia-reperfusion injury, pretreatment with NPY or (Leu³¹, Pro³⁴)−NPY was not able to prevent apoptosis or rescue RGCs. In conclusion, we found modulatory effects of NPY application that for the first time were detected at the level of RGCs. However, further studies are needed to evaluate whether NPY neuroprotective actions detected in retinal explants can be translated into animal models of retinal degenerative diseases.     

Synthesis of some pyrazolyl benzenesulfonamide derivatives as dual anti-inflammatory antimicrobial agents

Four series of pyrazolylbenzenesulfonamide derivatives were synthesized and evaluated for their anti-inflammatory activity using cotton pellet induced granuloma and carrageenan-induced rat paw edema bioassays. Moreover, COX-1 and COX-2 inhibitory activity, ulcerogenic effect and acute toxiCIT000y were also determined. Furthermore, the target compounds were screened for their in-vitro antimicrobial activity against Eischerichia coli, Staphylococcus aureus and Candida albicans. Compounds 4-(3-Phenyl-4-cyano-1H-pyrazol-1-yl)benzenesulfonamide 9a and 4-(3-Tolyl-4-cyano-1H-pyrazol-1-yl)benzenesulfonamide 9b were not only found to be the most active dual anti-inflammatory antimicrobial agents in the present study with good safety margin and minimal ulcerogenic effect but also exhibited good selective inhibitory activity towards COX-2. A docking pose for 9a and 9b separately in the active site of the human COX-2 enzyme was also obtained. Therefore, these compounds would represent a fruitful matrix for the development of dual anti-inflammatory antimicrobial candidates with remarkable COX-2 selectivity.