Femtosecond near-infrared laser pulses as a versatile non-invasive tool for intra-tissue nanoprocessing in plants without compromising viability

In this report, we describe a highly reproducible femtosecond near-infrared (NIR) laser-based nanoprocessing technique that can be used both for non-invasive intra-tissue nanodissection of plant cell walls as well as selective destruction of a single plastid or part thereof without compromising the viability of the cells. The ultra-precise intra-tissue nanoprocessing is achieved by the generation of high light intensity (10(12)W cm(-2)) by diffraction-limited focusing of the radiation of an NIR (lambda = 740 and 800 nm) femtosecond titanium-sapphire laser to a sub-femtolitre volume and subsequent highly localized instantaneous plasma formation. Following nanosurgery, electron microscopical analysis of the corresponding cellular target areas revealed clean non-staggering lesions across the cell wall with a cut width measuring less than 400 nm. To our knowledge, this is the smallest cut made non-invasively within a plant tissue. Further evidence, including two-photon imaging of chlorophyll fluorescence, revealed that a single target chloroplast or part thereof can be completely knocked out using intense ultra-fast NIR pulses without any visible deleterious effect on the adjacent plastids. The vitality of the cells after nanoprocessing has been ascertained by exclusion of propidium iodide from the cells as well as by the presence of cytoplasmic streaming. The potential applications of this technical advance include developmental biology applications, particularly studies addressing spatio-temporal control of ontogenetic events and cell-cell interactions, and gravitational biology applications.     

Lipid peroxidation and fluidity of plasma membranes from rat liver and Morris hepatoma 3924A

Plasma membranes isolated from the fast-growing, maximal-deviation, Morris hepatoma 3924A exhibit remarkable changes in lipid composition, lipid peroxidation and to some extent in the physical state with respect to rat liver plasmalemmas. A correlation appears to exit between the lower phospholipid: protein ratio, higher cholesterol: phospholipid ratio, lower rate of lipid peroxidation and decrease in fluidity in tumor plasma membranes.     

Ca2+-dependent ATPase associated with plasma membrane from a calcareous alga, Serraticardia maxima (Corallinaceae, Rhodophyta)

The plasma membrane was isolated from a calcareous red alga, Serraticardia maxima (Yendo) Silva (Corallinaceae), by aqueous two-phase partitioning. Its purity was examined with marker enzymes, Mg²⁺-dependent ATPase, inosine diphosphatase, cytochrome c oxidase and NADH-cytochrome c reductase, as well as the sensitivity of Mg²⁺-dependent ATPase to vanadate, azide and nitrate. The results showed that the isolated plasma membrane was purified enough to study its functions. Electron microscopic observations on thin tissue sections revealed that most vesicles of the isolated plasma membrane were stained by the plasma membrane specific stain, phosphotungstic acid-chromic acid. Mg²⁺- or Ca²⁺-dependent ATPases were associated with the plasma membrane. Ca²⁺-dependent ATPase was activated at physiological cytoplasmic concentrations of Ca²⁺ (0.1–10 μmol/L). However, calmodulin (0.5 μmol/L) did not affect its activity. The pH optimum was 8.0, in contrast to 7.0 for Mg²⁺-dependent ATPase. The isolated plasma membrane vesicles were mostly right side-out. To test for H⁺-translocation, right side-out vesicles were inverted; 27% of vesicles were inside-out after treatment with Triton X-100. The inside-out plasma membrane vesicles showed reduction of quinacrine fluorescence in the presence of 1 mmol/L ATP and 100 μmol/L Ca²⁺. The reduced fluorescence was recovered with the addition of 10 mmol/L NH₄Cl, or 5 μmol/L nigericin plus 50 mmol/L KCl. UTP and CTP substituted for ATP, but ADP did not. Ca²⁺-dependent ATPase might pump H⁺ out in the physiological state. The acidification by this pump might be coupled with alkalinization at the calcifying sites, which induces calcification.     

常压室温等离子体技术在微生物诱变中的应用进展

微生物是人类赖以生存的重要资源,为提高微生物的生产效率或者赋予其新的生物学功能,需要通过理化方法进行诱变或通过分子生物学技术对其进行定点突变。在目前的理化诱变方法中,常压室温等离子（atmospheric and room temperature plasma,ARTP）诱变技术具有操作简单、条件温和、安全性高、诱变快速等优点,成为倍受青睐的新方法。基于此,综述了ARTP诱变技术的原理及其在微生物诱变育种方面的应用,以期为选育性能优越的微生物菌种的诱变育种相关研究提供借鉴。     

Ionic and osmotic components of salt stress specifically modulate net ion fluxes from bean leaf mesophyll

Ionic mechanisms of salt stress perception were investigated by non‐invasive measurements of net H⁺, K⁺, Ca²⁺, Na⁺, and Cl^? fluxes from leaf mesophyll of broad bean (Vicia faba L.) plants using vibrating ion‐selective microelectrodes (the MIFE technique). Treatment with 90 m M NaCl led to a significant increase in the net K⁺ efflux and enhanced activity of the plasma membrane H⁺‐pump. Both these events were effectively prevented by high (10 m M ) Ca²⁺ concentrations in the bath. At the same time, no significant difference in the net Na⁺ flux has been found between low‐ and high‐calcium treatments. It is likely that plasma membrane K⁺ and H⁺ transporters, but not the VIC channels, play the key role in the amelioration of negative salt effects by Ca²⁺ in the bean mesophyll. Experiments with isotonic mannitol application showed that cell ionic responses to hyperosmotic treatment are highly stress‐specific. The most striking difference in response was shown by K⁺ fluxes, which varied from an increased net K⁺ efflux (NaCl treatment) to a net K⁺ influx (mannitol treatment). It is concluded that different ionic mechanisms are involved in the perception of the ‘ionic’ and ‘osmotic’ components of salt stress.     

Nonthermal plasma induces head and neck cancer cell death: the potential involvement of mitogen-activated protein kinase-dependent mitochondrial reactive oxygen species

Nonthermal plasma (NTP) is generated by ionization of neutral gas molecules, which results in a mixture of energy particles including electrons and ions. Recent progress in the understanding of NTP has led to its application in the treatment of various diseases, including cancer. However, the molecular mechanisms of NTP-induced cell death are unclear. The purpose of this study was to evaluate the molecular mechanism of NTP in the induction of apoptosis of head and neck cancer (HNC) cells. The effects of NTP on apoptosis were investigated using MTT, terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labeling, Annexin V assays, and western blot analysis. The cells were examined for production of reactive oxygen species (ROS) using DCFCA or MitoSOX staining, intracellular signaling, and an animal model. NTP reduced HNC cell viability in a dose-dependent manner and induced apoptosis. NTP resulted in alteration of mitochondrial membrane potential and accumulation of intracellular ROS generated from the mitochondria in HNC cells. Blockade of ROS production by N-acetyl-L-cysteine inhibited NTP-induced apoptosis. NTP led to the phosphorylation of c-JUN N-terminal kinase (JNK) and p38, but not extracellular-regulated kinase. Treatment with JNK and p38 inhibitors alleviated NTP-induced apoptosis via ROS generation. Taken together, these results show that NTP induced apoptosis of HNC cells by a mechanism involving MAPK-dependent mitochondrial ROS. NTP inhibited the growth of pre-established FaDu tumors in a nude mouse xenograft model and resulted in accumulation of intracellular ROS. In conclusion, NTP induced apoptosis in HNC cells through a novel mechanism involving MAPK-mediated mitochondrial ROS. These findings show the therapeutic potential of NTP in HNC.     

经尿道等离子前列腺剜除术与经尿道前列腺电切术治疗良性前列腺增生症的临床疗效对比研究

摘要 目的：对比经尿道等离子前列腺剜除术（TUKEP）与经尿道前列腺电切术（TURP）治疗良性前列腺增生症（BPH）的临床疗效。方法：回顾性分析我院2018年1月1日至2021年3月17日期间收治的200例BPH患者的临床资料。根据手术方式的不同将患者分为A组（n=83）和B组（n=117）,A组手术方式为TURP,B组手术方式为TUKEP,比较两组围术期指标,随访6个月,对比两组性功能、尿流动力学变化及并发症发生情况。结果：B组手术时间、术中出血量、尿管留置时间、住院时间、术后冲洗时间短于A组（P<0.05）。术后6个月,两组患者的最大尿流速（Qmax）、膀胱顺应性（BC）升高,剩余尿量（PVR）降低（P<0.05）,且B组患者的Qmax、BC高于A组,PVR低于A组（P<0.05）。术后6个月,两组患者的勃起功能评分表（IIEF-5）、射精功能评分表（CIPE-5）评分降低（P<0.05）,但B组、A组IIEF-5、CIPE-5评分组间对比无明显统计学差异（P>0.05）。B组的并发症发生率小于A组（P<0.05）。结论：TUKEP、TURP治疗BPH,疗效相当,TUKEP在缩短手术时间、尿管留置时间、术后冲洗时间、住院时间,降低术中出血量,减少并发症发生率,改善尿流动力学方面更有优势。     

亚甲蓝光化学法灭活新鲜冰冻血浆病毒及其效果评价

目的:探讨以亚甲蓝光化学法(MB-P)灭活新鲜冰冻血浆病毒及其效果。方法:选取2013年3月-2016年11月来青海省血液中心无偿献血者捐献的400 mL全血共1500份,分离新鲜冰冻血浆200 mL后,再分为两份即对照组和观察组各100 mL,各1500份。对照组不经灭活,实验组经MB-P病毒灭活。比较两组的纤维蛋白原(FIB)、总蛋白(TP)、凝血因子VⅢ(FVⅢ)及白细胞(WBC)的含量。再随机抽取500例接受了上述病毒灭活血浆输注的患者,观察其输血反应的发生情况。结果:观察组的FIB和TP含量及FVⅢⅠ活性比对照组有所降低,但差异不具有统计学意义(P0.05);观察组的WBC含量较对照组明显降低,且具有统计学差异(P0.05);FIB、TP、FVⅢⅠ的回收率的分别为85.43%、91.08%和80.49%,WBC的残留率为0.21%;500例接受了上述病毒灭活血浆输注的患者中,有1例出现发热症状。结论:经MB-P灭活病毒血浆的有效成分含量较高,符合国家相关标准,并且WBC含量和输血不良反应发生率均较低,满足临床安全输血的要求,但仍需严格要求血液收集和血浆病毒灭活的操作过程并严格掌握输血适应症,避免人为造成的输血风险。     

Imitational modeling of process of evolution: From organic macromolecules to protocell and animal cell

A dynamic imitational model of initial stages of cell evolution has been developed based on role of environmental calcium concentration. The model is designed from our hypothesis about the medium of the appearance of protocells, which could be potassium water reservoirs rather than sea salt water with its predominance of sodium salts. The necessary elements of the appearance of the protocells served organic molecules, code of their synthesis, and formation of macromolecules under favorable ion concentration in environment: a high K⁺ and Mg²⁺ and a low Na⁺ concentration. The model is based on an assumption that one of the first stages in evolution of life was the appearance in the potassium-magnesium water reservoirs of organic molecules capable for self-replication on the basis of genetic code and formation of protocell with the potassium cytoplasm. The model has demonstrated necessity of formation of cell envelope for development of the protocell. Replacement of the dominant cation in water reservoirs—potassium by sodium—required the appearance of ion-transporting devices in plasma membrane and their participation in adaptation of cells to environment. This stage of evolution was accompanied by the most important morphofunctional event—formation of the plasma membrane instead of cell envelope. The membrane provided the ion asymmetry in the cell (preservation of K⁺ in it) relatively to the sodium external medium for maintaining optimal intracellular medium. In the model system, predecessors of animal cells elaborated mechanism of maintenance of the potassium cytoplasm with the sodium counterion dominating in the environment.