Rb1基因第16内含子内21个碱基缺失1例

目地研究双眼视网膜母细胞瘤患者Rb1基因杂合性突变的分子生物学特性。方法应用PCR—SSCP直接测序技术检测双眼视网膜母细胞瘤患者白细胞DNA中Rb1基因杂合性突变。结果50例证实有Rb1基因杂合性突变的病例中有1例发生于第16内含子中可以用3种定位方法解释、具有相同序列的21个碱基缺失。结论这种极为少见的Rb1基因突变方式可能是由于破坏了正常拼接位点的结构而激活了“隐蔽拼接位点”，导致异常的Rb1基因mRNA产生或由此影响整个拼接过程。     

西洋参和胡萝卜体细胞融合培养杂种鉴定及愈伤组织中人参皂苷Rb_1含量分析

目的 :通过采用体细胞融合技术将西洋参基因转入胡萝卜中 ,为贵重中药、生长受地理环境等限制的中药扩大药源 ,利用杂种优势为培养适于大面积栽培且有效成分人参皂苷含量较高的优良杂交品种提供理论与实验依据。方法 :用PEG法对五加科植物西洋参与伞形科植物胡萝卜进行体细胞融合 ,通过同工酶进行初步杂种鉴定并用HPLC法测定西洋参和胡萝卜体细胞融合培养愈伤组织中人参皂苷Rb1含量。结果 :体细胞融合技术成功地获得了西洋参和胡萝卜体细胞杂交愈伤组织 ;同工酶分析是鉴定杂种的有效手段之一 ;在 10个杂交体愈伤组织中有 6个杂交体愈伤组织人参皂苷Rb1含量比未融合前西洋参愈伤组织中的含量高。     

人参皂甙单体Rb1对大鼠在体心脏收缩性能的影响

本文作者为进一步探讨人参皂甙单体Rb1对心肌收缩的影响,完善人参对心肌保护作用的系统性研究,拟在在体工作鼠心上,以心肌收缩性能为指标,对比观察Rb1对心肌收缩的影响.     

The role of Na+/K+ ATPase activity during low flow ischemia in preventing myocardial injury: A 31P, 23Na and 87Rb NMR spectroscopic study

An increase in intracellular Na⁺ during ischaemia has been associated with myocardial injury. In this study, we determined whether inhibition of Na⁺/K⁺ ATPase activity contributes to this increase and whether Na⁺/K⁺ ATPase activity can be maintained by provision of glucose to perfused rat hearts during low flow, 0.5 ml/min, ischemia. We used ³¹P NMR spectroscopy to determine changes in myocardial energetics and intracellular and extracellular volumes. ²³Na NMR spectroscopy, with DyTTHA^3- present as a shift reagent, was used to measure changes in intracellular Na⁺ and ⁸⁷Rb NMR spectroscopy was used to estimate Na⁺/K⁺ ATPase activity from Rb⁺ influx rates, Rb⁺ being an NMR-sensitive congener of K⁺. In hearts provided with 11 mM glucose throughout ischemia, glycolysis continued and ATP was twofold higher than in hearts without glucose. In the glucose-hearts, Rb⁺ influx rate was threefold higher, intracellular Na⁺ was fivefold lower at the end of ischemia and functional recovery during reperfusion was twofold higher. We propose that continuation of glycolysis throughout low flow ischemia allowed maintenance of sufficient Na⁺/K⁺ ATPase activity to prevent the increase in intracellular Na⁺ that would otherwise have led to myocardial injury.     

Rb基因在鼻咽癌中存在状态的研究

首次应用生物素－１４－ｄＵＴＰ标记Ｒｂ３．８ｋｂ探针，结合亲合素－碱性磷酸酶发光及自显影法，对Ｒｂ基因在鼻咽癌中存在状态进行了研究。杂交结果表明，３例正常胎儿鼻咽组织出现分子量为１２．８、１０．２、８．０、６．２、５．６、５．２及４．８ｋｂ的７条杂文区带，１１例鼻咽癌组织均发现有Ｒｂ基因缺失或失活，其中４例为５．６ｋｂ片段缺失，４例为４．８ｋｂ缺失并有２例伴５．６ｋｂ减弱，２例为１０．２ｋｂ缺失，１例为５．６及５．２ｋｂ片段显著减弱，说明在上述１１例鼻咽癌中均有Ｒｂ基因的改变。这种高频率的异常变化，提示Ｒｂ基因的缺失或失活，与鼻咽癌的发生有密切关系。     

Accumulation of p53 in response to adenovirus early region 1A sensitizes human cells to tumor necrosis factor alpha-induced apoptosis

Many tumor cells are resistant to tumor necrosis factor alpha (TNFalpha)-induced apoptosis. Adenovirus early region 1A (AdE1A) sensitizes the otherwise resistant cells to TNFalpha. AdE1A also stabilizes the p53 protein. The present study demonstrates a correlation between AdE1A-induced sensitization and stabilization of p53 in TNFalpha-induced apoptosis since the N-terminal and CR2 regions, the binding sites for CBP/p300, Rb and 26S proteasome regulatory components, are required for both these actions of AdE1A. TNFalpha does not induce apoptosis and AdE1A fails to sensitize TNFalpha cytotoxicity in p53-negative cells. However, introduction of exogenous p53 overcomes the cellular resistance to TNFalpha toxicity and enhances AdE1A sensitization, demonstrating that AdE1A sensitizes TNFalpha-induced apoptosis by its stabilization of p53. A proteasome inhibitor, lactacystin, enhances TNFalpha cytotoxicity in p53-positive and -negative cells, suggesting that accumulation of cellular proteins other than p53 might also regulate the cellular response to TNFalpha signaling.     

Cellular site of active K absorption in the guinea-pig distal colonic epithelium

 The mammalian distal colon, which is composed of different cell types, actively transports Na, K and Cl in absorptive and K and Cl in secretory directions. To further characterize the K absorption process and to identify the cells involved in K absorption, unidirectional Rb fluxes and luminal Rb uptake into different epithelial cell types were determined in isolated guinea-pig distal colon. Net Rb absorption (1.5–2.5 μmol·h^–1·cm^–2) was not influenced by inhibition of Na transport with amiloride or by incubating both sides of the epithelium with Na-free solutions, but was almost completely abolished by luminal ouabain, ethoxzolamide or by incubating both sides of the epithelium with Cl-free solutions. Luminal Rb uptake, blockable by luminal ouabain, preferentially occurred in columnar surface and neck cells, to a lesser extent in surface goblet cells and to an insignificant degree in lower crypt cells. Employing a luminal Rb-Ringer (5.4 mM Rb) the Rb concentration increased within 10 min in columnar surface and neck, surface goblet and lower crypt cells to 70, 32 and about 10 mmol·kg^–1 wet weight, respectively. The presence of 5.4 mM K in the luminal incubation solution reduced Rb uptake almost completely indicating a much higher acceptance of the luminal H-K-ATPase for K than for Rb. The increase in Na and decrease in K concentrations in surface and neck cells induced by luminal ouabain might indicate inhibition of the basolateral Na-K-ATPase or drastic enhancement of cellular Na uptake by the Na-H exchanger. Bilateral Na-free incubation did not alter Rb uptake, but bilateral Cl-free incubation drastically reduced it. Inhibition of net Rb absorption by ethoxzolamide and inhibition of both Rb absorption and Rb uptake by bilateral Cl-free incubation support the notion that cellular CO₂ hydration is a necessary prerequisite for K absorption and that HCO₃ leaves the cell via a Cl-HCO₃ exchanger. Since ouabain-inhibitable transepithelial Rb flux and luminal Rb uptake rate by surface and neck cells were about the same, Rb(K) absorption seems to be accomplished mainly by columnar surface cells. Received: 4 August 1997 / Received after revision: 12 November 1997 / Accepted: 4 December 1997     

Expression of cyclin D1, retinoblastoma gene protein, and p16 MTS1 protein in atypical adenomatous hyperplasia and adenocarcinoma of the lung

 To clarify the events leading to the disruption of cell growth control that occurs during the development of pulmonary adenocarcinoma (AC), we used immunohistochemistry to evaluate the expression of G1 cycle regulators, cyclin D1, Rb protein (pRb), and p16 MTS1 protein and the tumour proliferation marker, Ki 67, both in AC of the lung and in its precursor lesion, atypical adenomatous hyperplasia (AAH). The frequency of lesions with cyclin D1 overexpression was relatively high in AAH (47–89%), but was decreased in early AC (28%) and overt AC (35%). The loss of pRb expression was rare in both AAH (0–18%) and early AC (0%), and was infrequent even in overt AC (13%). The loss of p16 expression was also relatively infrequent in both the premalignant and the malignant lesions (11–25%). Our results suggest that overexpression of cyclin D1 is an early event and plays an important part in tumorigenesis in the case of lung AC. However, cyclin D1 overexpression is not required for the development and maintenance of a malignant phenotype. It is likely that some cyclin D1-independent pathways other than Rb and p16 abnormalities have an important role in the malignant transformation from AAH to early AC. Received: 8 July 1997 / 26 September 1997     

Cytogenetics of a new cytotype of African Mus (subgenus Nannomys) minutoides (Rodentia, Muridae) from Kenya: C- and G- banding and distribution of (TTAGGG) n telomeric sequences

We present the results of a cytogenetic study on Mus (Nannomys) minutoides from Kenya by means of C- and G- banding and in-situ fluorescence hybridization (FISH) to localize the telomeric sequences. The karyotype is characterized by the occurrence of several Rb chromosomes Rb(1.X), Rb(1.Y). Rb(2.17), Rb(3.13), Rb(4.10), Rb(5.11), Rb(6.7), Rb(8.12), not previously described for this species. This finding suggests a high level of chromosomal diversification, which means it is possible to consider this cytotype as a new, well-differentiated, chromosomal lineage within the subgenus. The C-banding of the metaphases illustrated conspicuous blocks of centromeric heterochromatin at the paracentromeric regions of all telocentric chromosomes. Centromeric heterochromatin is not visible on all biarmed chromosomes. Following hybridization with telomeric probes, bright interstitial telomeric sequence (ITS) fluorescence signals are evident at the pericentromeric area of all Rb chromosomes, with the exception of Rb(2.17). Considering the localization of the C-positive heterochromatin and of the telomeric sequences, the events leading to the Kenyan cytotype from an all-telocentric condition probably included two steps: first, fusion without loss of heterochromatin and pericentromeric telomeric sequences; second, the reduction of the C-positive satellite DNA followed by the amplification of telomeric sequences in the C-negative paracentromeric region of Rb chromosomes. The presence of a single Rb(2.17) without ITS indicates possible variations of this mechanism.     

Effects of selective α1 and α2 adrenoceptor active drugs on 86Rb+ efflux from pieces of rat parotid gland

Minute pieces of rat parotid gland were used in studies of adrenergic regulation of K+ efflux using ⁸⁶Rb+ as a probe for K+. Noradrenaline induced a concentration-dependent Rb+ efflux, whereas the β₁-selective agonist prenalterol was without effect. On the other hand, the β₂-selective drug, terbutaline, at high concentrations displayed a small enhancement of Rb+ -secretion. The selective α₁-adrenoceptor drug, phenylephrine, was as potent as noradrenaline, whereas the α₂-agonist clonidine had only a small effect. The noradrenaline-induced Rb+-efflux was effectively inhibited in the presence of prazosin, an α₁-blocker, whereas the α₂-antagonist, yohimbine, was roughly 50 times less potent. The results suggest that catecholamine-induced K+-secretion from the rat parotid gland is mediated via activation of post-synaptic α-adrenoceptors of the α₁-subtype.