Morphological heterogeneity and phenotypical instability versus metastatic stability in the murine tumor model ER 15-P

Summary At clinical presentation, the majority of malignant tumors are composed of multiple clonal subpopulations of tumor cells with different phenotypic characteristics. Using the experimental tumor model ER 15-P, a methylcholanthrene-induced pleomorphic sarcoma of the C57 Bl₆J mouse, we studied a system of long-term in vivo passages of this primary tumor for cell morphological changes, and alterations in the potential for spontaneous lung metastases. Transplants from the primary after the 4th, 20th, 40th and 80th i.m. passage (referred to as T4, T20, T40, and T80 respectively) together with their lung metastases were investigated by light microscopy, immunohistochemistry, and electron microscopy. In addition, the potential for metastasis to the lungs in each group was determined and compared with that of the parent T4 tumors. T4 tumors were mainly composed of spindle-shaped tumor cells with the ultrastructural features of fibroblasts and myofibroblasts, often arranged in a storiform or fasciculated growth pattern, and intermingled with tumor giant cells. Some small areas contained polygonal or rounded tumor cells, ultrastructurally undifferentiated, and sometimes arranged in a hemangiopericytoma-like growth pattern. Although electron-microscopical findings clearly demonstrated the mesenchymal origin of these tumor cells, immunostaining with a polyclonal antibody to vimentin was unspecific in all tumor cells and normal mouse tissue. Monoclonal antibodies to vimentin from different sources were completely negative in tumor cells and murine stromal components. In contrast, myofibroblast-like tumor cells showed immunohistochemically, a moderate to strong co-expression with monoclonal antibodies to desmin, muscle actin and -smooth muscle actin. On the basis of these morphological findings, the primary ER 15-P was classified as a pleomorphic myofibrosarcoma. The lung metastases of T4 tumors were mainly composed of undifferentiated round to polygonal tumor cells, while the number of desmin-positive, muscle- and -smooth muscle-actin-positive cells was reduced. The morphological features of T20 tumors and their lung metastases were the same as in T4, indicating a relative stability of the phenotype up to that stage. In contrast, T40 and T80 tumors and their lung metastases were found to contain almost exclusively undifferentiated tumor cells and many tumor giant cells. While fibroblast-like tumor cells were seen only occasionally, myofibroblast-like tumor cells had almost completely disappeared. The potential for lung metastases was nearly constant in all groups, suggesting metastatic stability. Obviously, the undifferentiated tumor cells of this model are associated with a higher metastatic potential.Abbreviations T4, T20, T40, T80 transplants from the primary tumor after the 4th, 20th etc. i.m. passage - MFH malignant fibrous histiocytoma     

Studies on the Level of Natural Antibodies Reactive with Various Tumor Cells during Urethane Carcinogenesis in BALB/c Mice

Serum from young normal BALB/c mice was found to contain IgM antibodies able tomediate complement-dependent lysis of certain syngeneic or allogeneic tumor target cells. The titer of such naturally occurring antitumor antibodies (NATA) was found to increase with aging.A longitudinal serological study comparing the cytotoxicity potential of NATA fromnormal and from urethan-treated BALB/c mice was performed. It was found that urethan-treated mice that did not develop primary lung-adenomas within the duration of the experi-ment had significantly lower NATA titers, against one out of 4 target cells assayed, than urethan-treated animals that developed lung adenomas. This difference was evident in two independent experiments. The results suggested that the lower NATA activity of the urethan-treated mice that did not develop tumors existed even before exposure to the carcinogenic insult. This raises the possibility that certain populations could be segregated according to their natural antibody profile into those individuals which will develop primary tumors within a certain period if exposed to a subthreshold amount of carcinogen, and those which will not.     

人胎盘CD133+细胞具有高增殖潜能集落形成细胞特性

目的 通过对人胎盘CD133⁺细胞群中高增殖潜能集落形成细胞(HPP-CFC)检测与生物学特性的分析，证明人胎盘存在早期造血干/祖细胞（HSPC）。 方法 采用机械法制备人胎盘组织（PT）单细胞悬液，用Histopaque-1007分离出单个核细胞(MNC)，经磁式分选（MACS）富集CD133⁺细胞，培养28 d后观察HPP-CFC集落形成能力，用流式细胞仪（FCM）对分选的细胞组份和HPP-CFC进行表型分析，实验全程用脐带血（UCB）作平行比较分析。 结果 培养28 d后，PT-CD133⁺与UCB-CD133⁺细胞组份分别扩增了266和362倍，前者低于后者（P<0.01）；PT-CD133⁺与UCB-CD133⁺细胞中HPP-CFC分别为(32.4±11.2)/5×10³、(17. 7±5.7)/5×10³，前者形成的HPP-CFC数量明显高于后者（P<0.01）；PT-CD133⁺、UCB-CD133⁺细胞培养至28 d时，除UCB-CD133⁺组的CD133⁺CD34^-亚群比例无明显改变外，CD133⁺CD34⁺、CD133^-CD34⁺和CD133⁺CD34^-（PT-CD133⁺组）亚型均比培养前减少。 结论 人胎盘组织CD133⁺细胞中存在HPP-CFC，说明胎盘CD133⁺细胞群中存在早期HSPC。     

A Field-Compatible Method for Interpolating Biopotentials

Mapping of bioelectric potentials over a given surface (e.g., the torso surface, the scalp) often requires interpolation of potentials into regions of missing data. Existing interpolation methods introduce significant errors when interpolating into large regions of high potential gradients, due mostly to their incompatibility with the properties of the three-dimensional (3D) potential field. In this paper, an interpolation method, inverse-forward (IF) interpolation, was developed to be consistent with Laplace's equation that governs the 3D field in the volume conductor bounded by the mapped surface. This method is evaluated in an experimental heart–torso preparation in the context of electrocardiographic body surface potential mapping. Results demonstrate that IF interpolation is able to recreate major potential features such as a potential minimum and high potential gradients within a large region of missing data. Other commonly used interpolation methods failed to reconstruct major potential features or preserve high potential gradients. An example of IF interpolation with patient data is provided to illustrate its applicability in the actual clinical setting. Application of IF interpolation in the context of noninvasive reconstruction of epicardial potentials (the inverse problem) is also examined. © 1998 Biomedical Engineering Society. PAC98: 8710+e, 0260Ed     

The sustained inward current in sino-atrial node cells of guinea-pig heart

 Single myocytes were dissociated from the sino-atrial (SA) node of guinea-pig hearts. Only a quite small fraction of the cell population showed spontaneous action potentials and these cells were characterized by the presence of the hyperpolarization-activated cation current I _f , the delayed rectifier K⁺ current I _K and the L-type Ca²⁺ current I _Ca,L as well as by the absence of both the transient outward current I _to and the inward rectifier K⁺ current I _K,1. After blocking I _f and I _K, depolarizing pulses from –80 mV revealed a large nicardipine-sensitive late current (NSLC). The NSLC was scarcely affected by decreasing extracellular [Ca²⁺] ([Ca²⁺]_o) from 1.8 to 0.1 mM, while it was decreased significantly by depleting [Na⁺]_o, differently from I _Ca,L. NSLC was blocked by nicardipine and was increased by Bay K 8644. NSLC was increased by isoprenaline and the additional application of acetylcholine reversed the increase of this current. We conclude that NSLC is largely composed of I _st described in the rabbit SA node pacemaker cells, and that I _st is unique for the pacemaker cells in mammalian SA node cells. Most of the quiescent cells showed neither I _f nor I _st. Received: 22 July 1996 / Received after revision: 30 September 1996 / Accepted: 9 October 1996     

Opioid peptides selective for mu- and delta-opiate receptors reduce calcium-dependent action potential duration by increasing potassium conductance

We suggest that both mu- and delta-opiate receptors on dorsal root ganglion neuron somata are coupled to voltage- and/or calcium-dependent potassium channels since opioid peptide decreases of calcium-dependent action potential duration were: (1) not associated with a change of resting membrane potential or conductance; (2) accompanied by an increase in action potential after-hyperpolarization, and (3) blocked by intracellular injection of the potassium channel blocker cesium [18]. In contrast, norepinephrine [4] and cadmium [9], which have been reported to act on voltage-dependent calcium rather than potassium channels, shortened action potential duration and decreased after-hyperpolarization amplitude, an action not blocked by intracellular iontophoresis of cesium.     

左氧氟沙星延长离体豚鼠乳头肌的动作电位时程

目的观察左氧氟沙星(LVFX)对豚鼠心肌动作电位的影响,并以司帕沙星(SPX)为阳性对照物比较二者对心肌的毒性作用。方法微电极记录豚鼠右心室乳头肌动作电位。结果刺激频率为1 Hz时,各浓度的SPX均显著性延长APD50和APD90,而LVFX浓度超过10μmol/L时明显延长APD50和APD90。结论LVFX和SPX均引起APD延长,前者效应较后者弱,仅在大剂量时有导致QT间期延长的潜在毒性。     

基于多分辨分析与连续小波变换提取和分析兔体感诱发电位

研究单次提取兔体感诱发电位,并定位和分析诱发电位波形成分。麻醉兔,以0.5Hz频率电脉冲刺激兔下肢隐神经,3764Hz采样率收集兔头皮电位。采用一维多分辨分析提取兔体感诱发电位,并用连续小波变换定位和分析诱发电位波形成分。单次诱发电位的小波变换与叠加平均诱发电位比较,表明Daubechies小波多分辨分析可以单次提取诱发电位。连续小波变换能够精确定位诱发电位中波形成分,并可采用连续小波变换分析诱发成分的频域特性。连续小波变换技术把一维时域信号投影到二维时频空间研究将成为医学信号处理的一个有用方法。     

Vasopressin induced rhythmic activity in rat basilar artery

The effects of vasopressin on membrane potential and tension were studied in isolated segments of basilar arteries from the University of Iowa colonies of normotensive inbred Kyoto-Wistar rats (WKY) and stroke-prone spontaneously hypertensive rats (SP-SHR). In the presence of vasopressin (0.01–0.3 IU/ml), basilar arteries from WKY, but not from SP-SHR, developed rhythmic contractions. These contractions were recorded in 13 of 14 WKY basilar arteries, were unaffected by pretreatment with 6-hydroxydopamine, and were characterized by 20–100 dyne oscillations in tension, occurring 1–3 cycles/min, and superimposed on the vasopressin-induced contraction (averaging 60 dynes at 0.01 IU/ml or 160 dynes at 0.3 IU/ml). However, resting membrane potentials were not different in SP-SHR vs. WKY at 37°C, and both strains showed about the same (11 mV) depolarization by 0.1 IU/ml of vasopressin. The rhythmic contractions were enhanced by K⁺-free solution, and abolished in the presence of high K⁺ solution (30 mM), suggesting that active Na⁺−K⁺ transport may be involved in modulating the rhythmic activity. These findings are consistent with the hypothesis that the vasopressin-induced rhythmic contractions in WKY basilar arteries are at least partly dependent on a reduced activity of electrogenic Na⁺−K⁺ active transport in WKY as compared to SP-SHR. This research was supported by Grant Nos. HL14388 and HL16328 from the National Institutes of Health. Dr. Rusch is the recipient of Postdoctoral Fellowship HL06907.     

A low-voltage activated,transient calcium current is responsible for the time-dependent depolarizing inward rectification of rat neocortical neurons in vitro

Intracellular recordings were obtained from rat neocortical neurons in vitro. The current-voltage-relationship of the neuronal membrane was investigated using current- and single-electrode-voltage-clamp techniques. Within the potential range up to 25 mV positive to the resting membrane potential (RMP: –75 to –80 mV) the steady state slope resistance increased with depolarization (i.e. steady state inward rectification in depolarizing direction). Replacement of extracellular NaCl with an equimolar amount of choline chloride resulted in the conversion of the steady state inward rectification to an outward rectification, suggesting the presence of a voltage-dependent, persistent sodium current which generated the steady state inward rectification of these neurons. Intracellularly injected outward current pulses with just subthreshold intensities elicited a transient depolarizing potential which invariably triggered the first action potential upon an increase in current strength. Single-electrode-voltage-clamp measurements reveled that this depolarizing potential was produced by a transient calcium current activated at membrane potentials 15–20 mV positive to the RMP and that this current was responsible for the time-dependent increase in the magnitude of the inward rectification in depolarizing direction in rat neocortical neurons. It may be that, together with the persistent sodium current, this calcium current regulates the excitability of these neurons via the adjustment of the action potential threshold.