Cell shape changes and transmembrane receptor uncoupling induced by tertiary amine local anesthetics

Tertiary amine local anesthetics (dibucaine, Tetracaine, procaine, etc.) modify cell morphology, concanavalin A (Con A)-mediated agglutinability and redistribution of Con A receptors. Con A agglutination of untransformed mouse 3T3 cells was enhanced at low concentrations of local anesthetics, and the dynamics of fluorescent-Con A indicated that ligand-induced clustering was increased in the presence of the drugs. In contast, these drugs inhibited Con A-induced receptor capping on mouse spleen cells. These effects can be duplicated by combinations of vinblastine (or colchicine) and cytochalasin B suggesting that local anesthetics act on microtubule cell surface receptor mobility and distribution. It is proposed that tertiary amine local anesthetics displace plasma membrane-bond Ca²⁺, resulting in disengagement of microfilament systems from the plasma membrane and increased cellular Ca²⁺ concentration to levels which disrupt microtubular organization. The possible involvement of cellular Ca²⁺ in cytoskeletal destruction by local anesthetics was investigated utilizing Ca²⁺-specific ionophores A23187 and X537A. In media containing Ca²⁺ and cytochalasin B these ionophores caused effects similar to tertiary amine local anesthetics.     

Mapping of neurons in the gravity receptor system of the octopus statocyst by iontophoretic cobalt staining

Summary The presence of uni-, bi- and multipolar neurons beneath the hair cell epithelium of the Octopus gravity receptor system has been demonstrated by iontophoretic cobalt staining. Counts give an average number of 1,940 neurons per macula. Whether the hair cells are primary of secondary sensory cells is discussed.This work was supported by grant Wo 160/3 of the Deutsche Forschungsgemeinschaft (DFG) to H.G.W. Thanks are due to the Director and staff of the Zoological Station in Naples for their hospitality and help     

Incorporation of methionine-[methyl-2H3] and mevalonic acid-[2-14C,(4R)-4-3H1] into Phycomyces blakesleeanus sterols

Ergosterol, episterol, 4α-methyl-5α-ergosta-8,24(28)-dien-3β-ol and 24-methylene-24,25-dihydrolanosterol, isolated from Phycomyces blakesleeanus grown in the presence of methionine-[methyl-²H₃], each contained two deuterium atoms; lanosterol, however, was unlabelled. The ¹⁴C:³H atomic ratio of the following sterols isolated from P. blakesleeanus grown in the presence of mevalonic acid-[2-¹⁴C,(4R)-4-³H₁], was: ergosterol, 5:3; episterol, 5:4; ergosta-5,7,24(28)-trien-3β-ol, 5:3; 4α-methyl-5α-ergosta-8,24(28)-dien-3β-ol, 5:4; 24-methylene-24,25-dihydrolanosterol, 6:5; lanosterol, 6:5. The significance of these results in terms of ergosterol biosynthesis is discussed.     

Biotransformation of pregnenolone - 7alpha-3H, progesterone - 7alpha-3H and dehydroepiandrosterone - 7alpha-3H by porcine fetal and maternal adrenal homogenate preparations.

The biotransformation of pregnenolone-7alpha-3H and of progesterone-7alpha-3H by porcine fetal and maternal adrenal homogenates at 56 and 112 days of pregnancy and of dehydroepiandrosterone-7alpha-3H by fetal adrenal homogenates has been investigated in vitro. Both pregnenolone-7alpha-3H and progesterone-7alpha-3H were metabolized extensively by maternal adrenal preparations, the principal radioactive metabolites isolated being cortisol, corticosterone, 11-deoxycortisol, deoxycorticosterone, 11beta-hydroxyprogesterone and androstenedione. In addition, 17alpha-hydroxyprogesterone, 20alpha-dihydroprogesterone and cortisone were formed from both substrates and 17alpha-hydroxypregnenolone and progesterone were formed from pregnenolone. Although essentially the same radioactive metabolites were isolated after incubation of fetal adrenal glands with pregnenolone-7alpha-3H or progesterone-7alpha-3H, a greater proportion of the radioactivity was associated with corticosteroids at 112 days of pregnancy than at 56 days. 11beta-Hydroxyandrostenedione and androstenedione were isolated and identified together with an unknown polar metabolite, after incubation of fetal adrenal tissue with dehydroepiandrosterone-7alpha-3H. These results are discussed in relation to feto-placental steroid biosynthesis and metabolism and the role of the fetal adrenal in the initiation of parturition in the pig.     

The ultrastructure of topical cutaneous calcinosis

Summary Mineralized plaques, which develop at the site of repeated subcutaneous injections of 100 g KMnO₄/0.2 H₂O in rats, were investigated by electron microscopy. The newly formed, delineated, white plaque tissue at the injection site consisted of numerous, mostly unaltered fibroblasts and collagen fibers, without participation of inflammatory cells. Some signs of cell injury were found in the center of the lesions. Numerous, irregularly distributed, small, mineralized foci were seen near the fibroblasts. These were formed by aggregation of small needle-like units (50 Å in diameter and 0.05–2.0 m long). These needle-shaped units were found either in vesicular, cell derived structures, considered to be shed cell processes or cell fragments, or on collagen fibers. Intramitochondrial deposits of such needle-like units were seen frequently. Fusion of smaller mineralized foci to larger plaques occured and then needle-shaped units were seen at the periphery of the electron-dense lesions. Hypotheses concerning the mechanism of experimental cutaneous calcinosis (soft tissue mineralization) are discussed and related to the findings of this study. Probable intracellular crystal deposition and mineralization in cell-derived structures were shown for the first time in topical cutaneous calcinosis.     

玉米联合固氮菌Kosakonia radicincitans GXGL-4A的分离鉴定与固氮特性研究

【目的】固氮微生物是生物固氮的主体,其菌种的选育更是生物固氮研究的基础。本实验室分离鉴定出一株新的固氮菌,并对其固氮相关活性进行初步研究。【方法】固氮菌采用Ashby无氮培养基进行分离纯化。通过形态特征分析、生理生化特征分析、16S r RNA基因序列分析和基因组扫描对固氮菌进行鉴定。利用靛酚蓝-分光光度法检测固氮菌的泌铵能力。固氮酶活性的测定使用乙炔还原法。【结果】从玉米根部分离到一株固氮菌株GXGL-4A,菌体短杆状,大小约为1.5μm×0.5μm,单个或常见2个菌体细胞串联在一起,革兰氏染色结果为阴性。16S r RNA基因序列分析结果表明该菌株与Kosakonia oryzae Fo8A1d 16S r RNA基因序列有95%的相似性,结合形态特征、生理生化特征和基因组扫描,将其命名为K.radicincitans GXGL-4A。对其进行固氮基因nif H的检测,经PCR扩增得到预期的296 bp条带;采用靛酚蓝-分光光度法,测得发酵液中铵态氮含量为2.5 mg/L,表明该菌株具有较好的泌铵能力。乙炔还原法测其固氮酶活性,以乙烯生产量表示,结果显示GXGL-4A菌株在无氮培养基上能够有效地还原乙炔,达到232.94 nmol C2H4/(m L·h)。【结论】菌株GXGL-4A是一株可较好地分泌铵的新的固氮菌,具有很好的研究价值。     

AvrRps4 effector family processing and recognition in lettuce

During pathogenesis, effector proteins are secreted from the pathogen to the host plant to provide virulence activity for invasion of the host. However, once the host plant recognizes one of the delivered effectors, effector‐triggered immunity activates a robust immune and hypersensitive response (HR). In planta, the effector AvrRps4 is processed into the N‐terminus (AvrRps4^N) and the C‐terminus (AvrRps4^C). AvrRps4^C is sufficient to trigger HR in turnip and activate AtRRS1/AtRPS4‐mediated immunity in Arabidopsis; on the other hand, AvrRps4^N induces HR in lettuce. Furthermore, AvrRps4^N‐mediated HR requires a conserved arginine at position 112 (R112), which is also important for full‐length AvrRps4 (AvrRps4^F) processing. Here, we show that effector processing and effector recognition in lettuce are uncoupled for the AvrRps4 family. In addition, we compared effector recognition by lettuce of AvrRps4 and its homologues, HopK1 and XopO. Interestingly, unlike for AvrRps4 and HopK1, mutation of the conserved R111 in XopO by itself was insufficient to abolish recognition. The combination of amino acid substitutions arginine 111 to leucine with glutamate 114 to lysine abolished the XopO‐mediated HR, suggesting that AvrRps4 family members have distinct structural requirements for perception by lettuce. Together, our results provide an insight into the processing and recognition of AvrRps4 and its homologues.     

Design and synthesis of thiazolidine-2,4-diones hybrids with 1,2-dihydroquinolones and 2-oxindoles as potential VEGFR-2 inhibitors: in-vitro anticancer evaluation and in-silico studies

A thiazolidine-2,4-dione nucleus was molecularly hybridised with the effective antitumor moieties; 2-oxo-1,2-dihydroquinoline and 2-oxoindoline to obtain new hybrids with potential activity against VEGFR-2. The cytotoxic effects of the synthesised derivatives against Caco-2, HepG-2, and MDA-MB-231 cell lines were investigated. Compound 12a was found to be the most potent candidate against the investigated cell lines with IC₅₀ values of 2, 10, and 40 µM, respectively. Furthermore, the synthesised derivatives were tested in vitro for their VEGFR-2 inhibitory activity showing strong inhibition. Moreover, an in vitro viability study against Vero non-cancerous cell line was investigated and the results reflected a high safety profile of all tested compounds. Compound 12a was further investigated for its apoptotic behaviour by assessing the gene expression of four genes (Bcl2, Bcl-xl, TGF, and Survivin). Molecular dynamic simulations authenticated the high affinity, accurate binding, and perfect dynamics of compound 12a against VEGFR-2.