Reduced responsiveness of touch (SA I) receptors in the cat following close arterial infusion of neomycin

Electrophysiological recordings were made from afferent nerve fibres supplying slowly adapting type I (SA I) cutaneous mechano-receptors in the cat during application of mechanical stimuli (20 mN constant force, 0.2 s rise time and 2 s plateau phase, every 30 s). Close arterial infusion of neomycin (2.5 mg/min for 5 min) strongly depressed the responsiveness of SA I receptors. A direct effect on the receptor is thought to be the mode of action.     

氨基糖苷类抗生素细菌内毒素的检测方法

参照ＵＳＰ（Ⅹ　Ⅻ）中鲎试法（ＬＴ法）最大有效稀释度（ＭＶＤＴ）公式，对６种氨基糖苷类抗生素进行ＬＴ法试验，以考察细菌内毒素检查法的可行性．实验结果表明，在ＭＶＤＴ公式给出的限度内合理稀释，ＬＴ法检查热原可行，与ＲＴ法相符率高．     

The aminoglycoside antibiotic, gentamicin, fails to block increases in miniature endplate potential frequency induced by the sulfhydryl reagent, N-ethylmaleimide, in low calcium solutions

N-ethylmaleimide (NEM) increases the frequency of miniature endplate potentials (MEPPs) at the adult rat hemidiaphragm. This sulfhydryl-alkylating agent produces comparable effects in the absence of added calcium (2 mM EGTA), suggesting that the drug releases calcium from internal stores, or promotes calcium-dependent release by depolarizing the nerve terminal or interacting more directly with the release mechanism. These increases in frequency are not blocked by the aminoglycoside antibiotic, gentamicin; although the latter agent reduces quantal content and the elevations in MEPP frequency induced by high pottasium solutions. The results suggest that gentamicin and NEM act at different sites at the presynaptic terminal, and that the aminoglycosides block voltage-dependent presynaptic calcium influx.     

Calcium antagonism and reversibility of gentamicin-induced loss of cochlear microphonics in the guinea pig

The perilymphatic space of the guinea pig cochlea was perfused with various concentrations of Ca²⁺, Mg²⁺ and gentamicin. Cochlear microphonic potentials (CM) were essentially stable when solutions contained 1 mM Ca²⁺ plus 2 mM Mg²⁺, 1 mM Ca²⁺ and no Mg²⁺ or when both Ca²⁺ and Mg²⁺ were omitted. In the absence of Ca²⁺, the presence of 2 mM Mg²⁺ or 1 mM EGTA markedly decreased CM.Addition of 3 mM gentamicin lowered CM, the magnitude of the effect depending on the concentration of Ca²⁺ present. Ten mM Ca²⁺ eliminated the action of the drug. In the absence of Ca²⁺, the effects of Mg²⁺ and gentamicin were additive.When the application of gentamicin was followed by perfusion with drug-free medium, suppression of CM by 3 mM drug could be reversed with 10 mM Ca²⁺ but not with 1 mM Ca²⁺. Loss of CM was irreversible when administration of 10 mM Ca²⁺ was delayed or when the drug concentration was increased to 10 mM.The results are consistent with our previous hypothesis of a biphasic mechanism of aminoglycoside toxicity: an initial action competitive with calcium ions and reversible and a second step, non-competitive and irreversible.     

Inhibition of aac(6′)-Ib-mediated amikacin resistance by nuclease-resistant external guide sequences in bacteria

Inhibition of bacterial gene expression by RNase P-directed cleavage is a promising strategy for the development of antibiotics and pharmacological agents that prevent expression of antibiotic resistance. The rise in multiresistant bacteria harboring AAC(6′)-Ib has seriously limited the effectiveness of amikacin and other aminoglycosides. We have recently shown that recombinant plasmids coding for external guide sequences (EGS), short antisense oligoribonucleotides (ORN) that elicit RNase P-mediated cleavage of a target mRNA, induce inhibition of expression of aac(6′)-Ib and concomitantly induce a significant decrease in the levels of resistance to amikacin. However, since ORN are rapidly degraded by nucleases, development of a viable RNase P-based antisense technology requires the design of nuclease-resistant RNA analog EGSs. We have assayed a variety of ORN analogs of which selected LNA/DNA co-oligomers elicited RNase P-mediated cleavage of mRNA in vitro. Although we found an ideal configuration of LNA/DNA residues, there seems not to be a correlation between number of LNA substitutions and level of activity. Exogenous administration of as low as 50 nM of an LNA/DNA co-oligomer to the hyperpermeable E. coli AS19 harboring the aac(6′)-Ib inhibited growth in the presence of amikacin. Our experiments strongly suggest an RNase P-mediated mechanism in the observed antisense effect.     

Disruption of gap junctions attenuates aminoglycoside-elicited renal tubular cell injury

BACKGROUND AND PURPOSE

Gap junctions play important roles in the regulation of cell phenotype and in determining cell survival after various insults. Here, we investigated the role of gap junctions in aminoglycoside-induced injury to renal tubular cells.

EXPERIMENTAL APPROACH

Two tubular epithelial cell lines NRK-E52 and LLC-PK1 were compared for gap junction protein expression and function by immunofluorescent staining, Western blot and dye transfer assay. Cell viability after exposure to aminoglycosides was evaluated by WST assay. Gap junctions were modulated by transfection of the gap junction protein, connexin 43 (Cx43), use of Cx43 siRNA and gap junction inhibitors.

KEY RESULTS

NRK-E52 cells expressed abundant Cx43 and were functionally coupled by gap junctional intercellular communication (GJIC). Exposure of NRK-E52 cells to aminoglycosides, G418 and hygromycin, increased Cx43 phosphorylation and GJIC. The aminoglycosides also decreased cell viability that was prevented by gap junction inhibitors and Cx43 siRNA. LLC-PK1 cells were gap junction-deficient and resistant to aminoglycoside-induced cytotoxicity. Over-expression of a wild-type Cx43 converted LLC-PK1 cells to a drug-sensitive phenotype. The gap junction inhibitor α-glycyrrhetinic acid (α-GA) activated Akt in NRK-E52 cells. Inhibition of the Akt pathway enhanced cell toxicity to G418 and abolished the protective effects of α-GA. In addition, gentamycin-elicited cytotoxicity in NRK-E52 cells was also significantly attenuated by α-GA.

CONCLUSION AND IMPLICATIONS

Gap junctions contributed to the cytotoxic effects of aminoglycosides. Modulation of gap junctions could be a promising approach for prevention and treatment of aminoglycoside-induced renal tubular cell injury.     

产ESBLs大肠埃希菌的氨基糖苷抗菌药物耐药性及氨基糖苷类钝化酶基因检测

目的:了解地区产超广谱β-内酰胺酶(ESBLs)大肠埃希菌对氨基糖苷抗生素的耐药情况及氨基糖苷类钝化酶基因分布.方法:采用Kirby-Bauer(K-B)纸片法检测38株产 ESBLs大肠埃希菌对庆大霉素、阿米卡星、妥布霉素、奈替米星4种氨基糖苷类抗菌药物的耐药情况;并应用PCR方法检测这38株菌6种氨基糖苷钝化酶基因aac(3)-Ⅰ、aac(3)-Ⅱ、aac(6′)-Ⅱ、aac(6′)-Ib-cr、ant(2″)-Ⅰ、ant(3″)-Ⅰ.结果:38株产ESBLs大肠埃希菌对4种氨基糖苷类抗菌药物的耐药率分别为阿米卡星28.9%,奈替米星39.5%,庆大霉素76.3%,妥布霉素76.3%;aac(3)-Ⅱ、aac(6′)-Ib-cr、ant(3″)-Ⅰ、ant(2″)-Ⅰ阳性率分别为63.2%,36.8%,10.5%,2.6%.未检测出aac(3)-Ⅰ与aac(6′)-Ⅱ基因阳性菌株.结论:产ESBLs大肠埃希菌的氨基糖苷类钝化酶基因以aac(3)-Ⅱ、aac(6′)-Ib-cr基因为主,氨基糖苷类钝化酶基因与氨基糖苷类药物耐药性有一定的联系.临床抗感染过程中应该注重耐药机制的研究.     

Clinical and molecular epidemiology of hospital Enterococcus faecalis isolates in eastern France

Objective: To report on the occurrence of Enterococcus faecalis hospital isolates obtained during 1 year in hospitals in the Franche-Comté region of France.
Methods: Clinical isolates of E. faecalis of different antibiotic susceptibility phenotypes from hospitalized patients were characterized by pulsed-field gel electrophoresis. Patients with positive cultures were investigated by three case-control studies to identify risk factors for colonization/infection.
Results: The crude incidence of colonization/infection was 2.37%, and 4-day and 7-day colonization rates after admission were 10.0% and 6.36%, respectively. The rates of high-level resistance to kanamycin (HLKR) and to gentamicin (HLGR) were 47.1% and 7.1%, respectively. No isolate was resistant to glycopeptides or produced β-lactamase. The 209 hospital isolates obtained during the study yielded 98 major DNA patterns, of which two were major epidemic patterns including HLKR isolates. No single factor was significantly associated with colonization/infection by HLKR isolates. The length of hospitalization before isolation was associated with colonization by HLGR isolates.
Conclusions: The isolation frequency of E. faecalis strains with acquired resistance to aminoglycoside antibiotics, and the wide dissemination of resistant strains with characteristics that allow them to persist and spread, argue for further large prospective surveys of clinical isolates of E. faecalis in hospitals.     

庆大霉素对内侧橄榄耳蜗传出神经毒性作用的形态学观察

目的观察庆大霉素慢性耳中毒前后蒙古沙鼠内侧橄榄耳蜗（ｍｅｄｉａｌｏｌｉｖｏｃｏｃｈｌｅａｒ,ＭＯＣ）传出神经的形态改变及其与毛细胞损害的关系,以探讨ＭＯＣ传出神经在氨基糖甙类抗生素慢性耳中毒中的重要性。方法采用改良的乙酰胆碱酯酶组化染色和甲苯胺蓝苏木素染色法,全耳蜗铺片观察健康对照组和庆大霉素组耳蜗ＭＯＣ传出神经和外毛细胞（ｏｕｔｅｒｈａｉｒｃｅｌ,ＯＨＣ）的分布特征,并测量耳蜗ＭＯＣ传出神经纤维、末梢及ＯＨＣ的数量。结果停药后３周出现耳蜗ＭＯＣ纤维和末梢损害,损害程度随观察时间延长而加重,１１周最明显,且损害部位主要在耳蜗底回,与毛细胞损害的特征一致。结论耳蜗ＭＯＣ传出神经可能在庆大霉素慢性耳中毒时ＯＨＣ的损害中起重要作用。