A new chronological survival assay in mammalian cell culture

c-Abl is activated by oxidative stress but its precise function in cell response to this stress is elusive. Studies of c-Abl-/- osteoblasts revealed that c-Abl played a negative role in the induction of peroxiredoxin I (Prx I, Prdx I), an anti-oxidant protein with tumor suppression activity, but it provided a protective role against oxidative stress. In contrast, Atm, a signaling molecule that interacts with c-Abl and is required for c-Abl activation, served a totally different function. The significance of these findings is discussed here in the context of aging and tumorigenesis and their links to reactive oxygen species.     

Metal ion and inter-domain interactions as functional networks in E. coli topoisomerase I

Escherichia coli topoisomerase I (EcTopoI) is a type IA bacterial topoisomerase which is receiving large attention due to its potential application as novel target for antibacterial therapeutics. Nevertheless, a detailed knowledge of its mechanism of action at molecular level is to some extent lacking. This is partly due to the requirement of several factors (metal ions, nucleic acid) to the proper progress of the enzyme catalytic cycle. Additionally, each of them can differently affect the protein structure.     

三阴性乳腺癌中Ki-67 的表达及其临床病理意义

目的:探讨三阴性乳腺癌中Ki-67的表达及其临床病理意义。方法:回顾性分析2010年11月1日至2015年12月31日辽宁省肿瘤医院收治的110例行乳腺癌根治术的三阴性乳腺癌患者的临床病理资料,检测其乳腺癌组织中Ki-67的表达,并分析其与患者年龄、绝经情况、组织学分型、临床分期、淋巴结转移、肿瘤大小等临床病理资料的关系。进一步采用Kaplan-Meier生存分析法绘制生存曲线,比较高表达Ki-67和低表达Ki-67患者的3年、5年生存率。结果:Ki-67在三阴性乳腺癌患者50岁以下和50岁以上相比较,差异具有统计学意义(P0.05);Ki-67在三阴性乳腺癌患者绝经情况、组织学分型、临床分期、淋巴结转移、肿瘤大小的表达相比较,差异不具有统计学意义(P0.05);高表达Ki-67组3年生存率为79.49%,5年生存率为30.77%;低Ki-67组的3年生存率为85.92%,5年生存率为46.48%;两组3年生存率相比较差异不具有统计学意义(P0.05);两组5年生存率相比较差异具有统计学意义(P0.05);两组总生存时间相比较,差异具有统计学意义(P0.05)。结论:三阴性乳腺癌中Ki-67呈高表达,可作为乳腺癌预后不良的危险因素。     

Generation and preclinical characterization of an antibody specific for SEMA4D

Semaphorin 4D (SEMA4D or CD100) is a member of the semaphorin family of proteins and an important mediator of the movement and differentiation of multiple cell types, including those of the immune, vascular, and nervous systems. Blocking the binding of SEMA4D to its receptors can result in physiologic changes that may have implications in cancer, autoimmune, and neurological disease. To study the effects of blocking SEMA4D, we generated, in SEMA4D-deficient mice, a panel of SEMA4D-specific hybridomas that react with murine, primate, and human SEMA4D. Utilizing the complementarity-determining regions from one of these hybridomas (mAb 67-2), we generated VX15/2503, a humanized IgG4 monoclonal antibody that is currently in clinical development for the potential treatment of various malignancies and neurodegenerative disorders, including multiple sclerosis and Huntington's disease. This work describes the generation and characterization of VX15/2503, including in vitro functional testing, epitope mapping, and an in vivo demonstration of efficacy in an animal model of rheumatoid arthritis.     

结直肠癌组织RPN11与Ki67的表达及其临床病理学意义

目的观察去泛素化酶RPN11和增殖相关核标记物Ki67在结直肠癌组织中的表达,研究其与结直肠癌肿瘤细胞增殖的相关性及与结直肠癌临床病理特征的关系。方法采用免疫组织化学SABC法检测56例结直癌组织及20例癌旁正常组织中的RPN11和Ki67表达,结合临床病理学资料进行统计分析。结果免疫组织化学染色显示:RPN11及Ki67在结直肠癌组织的阳性表达率明显高于正常结直肠组织;RPN11和Ki67的表达均与肿瘤分化程度、TNM分期、转移有关,而与性别、年龄无明显相关;RPN11与Ki67的表达呈正相关。结论RPN11和Ki67可能共同参与结直肠癌肿瘤细胞的增殖调控,并促进结直肠癌的发生发展以及浸润转移。     

Neuronal nicotinic receptor deficits in Alzheimer patients with the Swedish amyloid precursor protein 670/671 mutation

The influence of beta-amyloid on cholinergic neurotransmission was studied by measuring alterations in nicotinic acetylcholine receptors (nAChRs) in autopsy brain tissue from subjects carrying the Swedish amyloid precursor protein (APP) 670/671 mutation. Significant reductions in numbers of nAChRs were observed in various cortical regions of the Swedish 670/671 APP mutation family subjects (-73 to -87%) as well as in sporadic Alzheimer's disease (AD) cases (-37 to -57%) using the nicotinic agonists [3H]epibatidine and [3H]nicotine, which bind with high affinity to both alpha3 and alpha4 and to alpha4 nAChR subtypes, respectively. Saturation binding studies with [3H]epibatidine revealed two binding sites in the parietal cortex of AD subjects and controls. A significant decrease in Bmax (-82%) for the high-affinity site was observed in APP 670/671 subjects with no change in K(D) compared with controls (0.018 nM APP 670/671; 0.036 nM control). The highest load of neuronal plaques (NPs) was observed in the parietal cortex of APP 670/671 brains, whereas the number of [3H]nicotine binding sites was less impaired compared with other cortical brain regions. Except for a positive significant correlation between the number of [3H]nicotine binding sites and number of NPs in the parietal cortex, no strict correlation was observed between nAChR deficits and the presence of NPs and neurofibrillary tangles, suggesting that these different processes may be closely related but not strictly dependent on each other.     

Effects induced by mono- and divalent cations on protein regions responsible for thermal adaptation in β-glycosidase from<Emphasis Type="Italic"> Sulfolobus solfataricus</Emphasis>

The perturbation induced by mono- and divalent cations on the thermophilicity and thermostability of Solfolobus solfataricus -glycosidase, a hyperthermophilic tetrameric enzyme, has been investigated by spectroscopic and computational simulation methods to ascertain the Hofmeister effects on two strategic protein regions identified previously. Specifically, (1) an extra segment (83–124), present only in the sequence of hyperthermophilic glycosidases and recognized as an important thermostability determinant for the enzyme structure; and (2) a restricted area of the subunit interface responsible for the quaternary structure maintenance. Mono- and divalent cations inhibit to a different extent the -glycosidase activity, whose kinetic constants show an apparent competitive inhibition of the catalytic process that reflects the Hofmeister order. The thermostability is also affected by the nature and charge of the cations, reaching maximal effects for the case of Mg²⁺. Fourier transform infrared spectroscopy has revealed very small changes in the protein secondary structure in the presence of the investigated cations at 20 °C, while large effects on the protein melting temperatures are observed. Computational analysis of the enzyme structure has identified negative patches on the accessible surface of the two identified regions. Following the Hofmeister series, cations weaken the existing electrostatic network that links the extra segment to the remaining protein matrix. In particular, the perturbing action of cations could involve the ionic pair interactions E107–R245 and E109–R185, thus leading to a local destructuring of the extra segment as a possible starting event for thermal destabilization. A detailed investigation of the electrostatic network at the A–C intermolecular interface of Sgly after energy minimization suggests that cations could cause a strong attenuation of the ion pair interactions E474–K72 and D473–R402, with consequent partial dissociation of the tetrameric structure.Abbreviations Amide I amide I band in a ²H₂O medium - EM energy minimization - ONPG o-nitrophenyl--d-galactopyranoside - Sgly Escherichia coli expressed Sulfolobus solfataricus -glycosidase     

N-Acetylcysteine ameliorates lipopolysaccharide-induced organ damage in conscious rats

Lipopolysaccharide is strongly associated with septic shock, leading to multiple organ failure. It can activate monocytes and macrophages to release proinflammatory mediators such as tumor necrosis factor- (TNF-), interleukin-1 (IL-1), and nitric oxide (NO). The present experiments were designed to induce endotoxin shock by an intravenous injection ofKlebsiella pneumoniae lipopolysaccharide (LPS, 10 mg/kg) in conscious rats. Arterial pressure and heart rate (HR) were continuously monitored for 48 h after LPS administration. N-Acetyl-cysteine was used to study its effects on organ damage. Biochemical substances were measured to reflect organ functions. Biochemical factors included blood urea nitrogen (BUN), creatinine (Cre), lactic dehydrogenase (LDH), creatine phosphokinase (CPK), aspartate transferase (GOT), alanine transferase (GPT), TNF-, IL-1, methyl guanidine (MG), and nitrites/nitrates. LPS caused significant increases in blood BUN, Cre, LDH, CPK, GOT, GPT, TNF-, IL-1, MG levels, and HR, as well as a decrease in mean arterial pressure and an elevation of nitrites/nitrates. N-Acetylcysteine suppressed the release of TNF-, IL-1, and MG, but enhanced NO production. These actions ameliorate LPS-induced organ damage in conscious rats. The beneficial effects may suggest a potential chemopreventive effect of this compound in sepsis prevention and treatment.