The ageing lens and cataract: a model of normal and pathological ageing

Cataract is a visible opacity in the lens substance, which, when located on the visual axis, leads to visual loss. Age-related cataract is a cause of blindness on a global scale involving genetic and environmental influences. With ageing, lens proteins undergo non-enzymatic, post-translational modification and the accumulation of fluorescent chromophores, increasing susceptibility to oxidation and cross-linking and increased light-scatter. Because the human lens grows throughout life, the lens core is exposed for a longer period to such influences and the risk of oxidative damage increases in the fourth decade when a barrier to the transport of glutathione forms around the lens nucleus. Consequently, as the lens ages, its transparency falls and the nucleus becomes more rigid, resisting the change in shape necessary for accommodation. This is the basis of presbyopia. In some individuals, the steady accumulation of chromophores and complex, insoluble crystallin aggregates in the lens nucleus leads to the formation of a brown nuclear cataract. The process is homogeneous and the affected lens fibres retain their gross morphology. Cortical opacities are due to changes in membrane permeability and enzyme function and shear-stress damage to lens fibres with continued accommodative effort. Unlike nuclear cataract, progression is intermittent, stepwise and non-uniform.     

Cross- and auto-correlation in early vision

Neurons that respond selectively to the orientation of visual stimuli were discovered in V1 more than 50 years ago, but it is still not fully understood how or why this is brought about. We report experiments planned to show whether human observers use cross-correlation or auto-correlation to detect oriented streaks in arrays of randomly positioned dots, expecting that this would help us to understand what David Marr called the 'computational goal' of V1. The streaks were generated by two different methods: either by sinusoidal spatial modulation of the local mean dot density, or by introducing coherent pairs of dots to create moiré patterns, as Leon Glass did. A wide range of dot numbers was used in the randomly positioned arrays, because dot density affects cross- and auto-correlation differently, enabling us to infer which method was used. This difference stems from the fact that the cross-correlation task is limited by random fluctuations in the local mean density of individual dots in the noisy array, whereas the auto-correlation task is limited by fluctuations in the numbers of randomly occurring spurious pairs having the same separation and orientation as the deliberately introduced coherent pairs. After developing a new method using graded dot luminances, we were able to extend the range of dot densities that could be used by a large factor, and convincing results were obtained indicating that the streaks generated by amplitude modulation were discriminated by cross-correlation, while those generated as moiré patterns were discriminated by auto-correlation. Though our current results only apply to orientation selectivity, it is important to know that early vision can do more than simple filtering, for evaluating auto-correlations opens the way to more interesting possibilities, such as the detection of symmetries and suspicious coincidences.     

促泌素在肾上腺原发肿瘤中的表达及其临床意义

目的比较促泌素(secretagogin,SCGN)与传统神经内分泌标记物在肾上腺原发肿瘤中的表达差异。方法收集肾上腺原发肿瘤手术标本共37例,其中包括18例皮质腺瘤、3例皮质腺癌、16例嗜铬细胞瘤。同时选取5例正常肾上腺组织,5例肾透明细胞癌作为对照。所有标本均使用SCGN、PGP9.5、CD56、NSE、Syn及CgA进行免疫组织化学SP法染色。结果SCGN在全部5例正常肾上腺皮质均有表达,而在髓质不表达(P<0.01),其中在皮质的表达明显高于PGP9.5和CgA的表达(P均<0.01);全部18例皮质腺瘤均表达SCGN,且明显高于NSE(P<0.05)、PGP9.5和CgA(P均<0.01);肾上腺嗜铬细胞瘤中SCGN的阳性表达率仅为18.8%(3/16),明显低于其它标记物(P均<0.01)。SCGN在皮质腺瘤(18/18)中的表达明显高于嗜铬细胞瘤(3/16)(P<0.01),而PGP9.5和CgA在嗜铬细胞瘤(15/16,16/16)中的表达明显高于皮质腺瘤(3/18,1/18)(P均<0.01);CD56、NSE和Syn在皮质腺瘤、皮质腺癌和嗜铬细胞瘤中均有高表达,但两两组间比较均无统计学差异(P均>0.05)。SCGN在全部5例肾透明细胞癌中均不表达。结论SCGN对肾上腺皮质腺瘤有较高敏感性,其与嗜铬细胞瘤的标记物CgA和PGP9.5联合可在两者的诊断和鉴别诊断中发挥重要作用。     

The RNA binding protein Musashi1 regulates apoptosis,gene expression and stress granule formation in urothelial carcinoma cells

The RNA‐binding protein Musashi1 (MSI1) is a marker of progenitor cells in the nervous system functioning as a translational repressor. We detected MSI1 mRNA in several bladder carcinoma cell lines, but not in cultured normal uroepithelial cells, whereas the paralogous MSI2 gene was broadly expressed. Knockdown of MSI1 expression by siRNA induced apoptosis and a severe decline in cell numbers in 5637 bladder carcinoma cells. Microarray analysis of gene expression changes after MSI1 knockdown significantly up‐regulated 735 genes, but down‐regulated only 31. Up‐regulated mRNAs contained a highly significantly greater number and density of Musashi binding sites. Therefore, a much larger set of mRNAs may be regulated by Musashi1, which may affect not only their translation, but also their turnover. The study confirmed p21^CIP1 and Numb proteins as targets of Musashi1, suggesting additionally p27^KIP1 in cell‐cycle regulation and Jagged‐1 in Notch signalling. A significant number of up‐regulated genes encoded components of stress granules (SGs), an organelle involved in translational regulation and mRNA turnover, and impacting on apoptosis. Accordingly, heat shock induced SG formation was augmented by Musashi1 down‐regulation. Our data show that ectopic MSI1 expression may contribute to tumorigenesis in selected bladder cancers through multiple mechanisms and reveal a previously unrecognized function of Musashi1 in the regulation of SG formation.     

Effects of testosterone on nuclear ribonucleoprotein components of prostate epithelial cells

Summary— The changes of the nuclear components caused by castration and testosterone injection were studied in epithelial cells of the ventral prostate of the rat. Castration drastically diminishes the nuclear and nucleolar volume, as well as the fraction of the nuclear volume occupied by non-nucleolar ribonucleoprotein (RNP) fibrils. However, in castrated animals the frequency of perichromatin granules (PCG) is 79% higher than in controls. Testosterone injection causes a reduction of the number of PCG to 33% of the castrated level in 15 min, and increases the non-nucleolar RNP fibrils. Other parameters such as nuclear and nucleolar volume and the relative volume of the compact chromatin present only small changes in a period of 2 h following the hormone administration. High resolution quantitative autoradiography demonstrates that the transportation of previously synthesized RNA increases steeper than the RNA synthesis. All these effects are similar to those caused by ovariectomy and estradiol injection on the nuclear structures of endometrial epithelial cells. These similarities and other observations suggest that PCGs contain mRNA, of a few genes, stored in the nucleus by a restriction of its transportation to the cytoplasm.     

Acetylcholine Release Induced by the Volatile Anesthetic Sevoflurane in Rat Brain Cortical Slices

1. We have investigated the effect of the volatile anesthetic sevoflurane on acetylcholine (ACh) release from rat brain cortical slices. 2. The release of [3H]-ACh into the incubation fluid was studied after labeling the tissue ACh with [methyl-3H]-choline chloride. 3. We observed that sevoflurane induced an increase on the release of ACh that was dependent on incubation time and anesthetic concentration. The sevoflurane-induced ACh release was not blocked by tetrodotoxin (TTX) and therefore was independent of sodium channels. In addition, the sevoflurane effect was not blocked by ethylene glycol-bis(beta-aminoethyl ether (EGTA) or cadmium (Cd2+), thus independent of extracellular calcium. 4. The sevoflurane-induced ACh release was inhibited by 1,2-bis (2-aminophenoxy) ethane-N,N,N',N'-tetra-acetic acid (BAPTA-AM), suggesting the involvement of intracellular calcium-sensitive stores in the process. Dantrolene, an inhibitor of ryanodine receptors, had no effect but 2-aminoethoxydiphenylborate (2-APB), a membrane-permeable inhibitor of inositol 1,4,5-triphosphate receptor inhibited the sevoflurane-induced release of ACh. 5. It is concluded that sevoflurane-induced release of ACh in brain cortical slices involves the mobilization of calcium from IP3-sensitive calcium stores.     

Novel regulation of Na, K-ATPase by Src tyrosine kinases in cortical neurons

The Na+, K+-ATPase or Na+, K+-pump plays a critical role in ion homeostasis and many cellular events. The Na+, K+-pump activity is regulated by serine/threonine phosphorylation, the role of tyrosine kinases in the regulation, however, is obscure. We now present novel evidence showing that tyrosine phosphorylation activates the Na+, K+-pump in cortical neurons. The electrogenic activity of the Na+, K+-pump was measured using whole-cell voltage clamp. A tonic activity was revealed by an inward current induced by the specific inhibitor ouabain or strophanthidin; an outward current due to activation of the pump was triggered by raising extracellular K+. The inward and outward currents were attenuated by the tyrosine kinase inhibitor genistein, herbimycin A, or lavendustin A, while blocking tyrosine phosphatases increased the pump current. Down-regulation of the pump current was also seen with the Src inhibitor PP1 and intracellularly applied anti-Lyn or anti-Yes antibody. Consistently, intracellular application of Lyn kinase up-regulated the pump current. Immunoprecipitation and western blotting showed tyrosine phosphorylation and a direct interaction between Lyn and the alpha3 subunit of the Na+, K+-pump. The tyrosine phosphorylation of the alpha3 subunit was reduced by serum deprivation. These data suggest that the Na+, K+-ATPase activity in central neurons is regulated by specific Src tyrosine kinases via a protein-protein mechanism and may play a role in apoptosis.     

Fine structure of the midgut and Malpighian papillae in Campodea (Monocampa) quilisi Silvestri, 1932 (Hexapoda, Diplura) with special reference to the metal composition and physiological significance of midgut intracellular electron-dense granules

The fine structure of the midgut and the Malpighian papillae in Campodea (Monocampa) quilisi Silvestri, 1932 (Hexapoda, Diplura) specimens was described. We observed the presence of electron-dense granules (EDGs) in the midgut epithelial cells, similar in genesis, structure and aspect to the type A spherocrystals described in the midgut epithelium of Collembola and Diplopoda. Energy-dispersive X-ray microanalysis was used to detect the chemical composition of the granules and to relate it to the concentrations of some potential toxic heavy metals (Pb, Cu, Zn) in soil and litter. Chemical composition of the granules seems strongly influenced by the presence and bioavailability of heavy metals in the external environment. Specimens from a contaminated abandoned mining and smelting area (Colline Metallifere, southern Tuscany) were able to accumulate Fe, Mn, Zn, Pb and Cu in their midgut EDGs. In addition, we observed that C. (M.) quilisi was able to excrete the metal-containing granules into the external medium by the moulting of the intestinal epithelium. This confirms that the process of ionic retention of midgut cells is particularly significant in animals lacking Malpighian tubules.     

Xenopus galectin-VIIa binds N-glycans of members of the cortical granule lectin family (xCGL and xCGL2)

We have identified members of the Xenopus cortical granule lectin (xCGL) family as candidate target glycoproteins of Xenopus galectin-VIIa (xgalectin-VIIa) in Xenopus embryos. In addition to the original xCGL, we also identified a novel member of the xCGL family, xCGL2. Expression of the mRNAs of xCGL and xCGL2, as well as that of xgalectin-VIIa, was observed throughout early embryogenesis. Two and three potential N-glycosylation sites were deduced from the amino acid sequences of xCGL and xCGL2, respectively, and xgalectin-VIIa recognizes N-glycans linked to a common site in xCGL and xCGL2 and also recognizes N-glycans linked to a site specific to xCGL2. However, interaction between xgalectin-Ia and xCGLs was not detectable. We also obtained consistent results on surface plasmon resonance analysis involving xCGLs as ligands and xgalectins as analytes. The Kd value of the interaction between xgalectin-VIIa and xCGLs was calculated to be 35.9 nM. The structures of the N-glycans of xCGLs, which were recognized by xgalectin-VIIa, were analyzed by the two-dimensional sugar map method, and three kinds of N-acetyllactosamine type, biantennary N-glycans were identified as the major neutral N-glycans. The binding specificity of oligosaccharides for xgalectin-VIIa was analyzed by frontal affinity chromatography (FAC). The oligosaccharide specificity pattern of xgalectin-VIIa was similar to that of the human homolog galectin-3, and it was also shown that the N-acetyllactosamine type, biantennary N-glycans exhibit high affinity for xgalectin-VIIa (Kd = 11 microM). These results suggest that xgalectin-VIIa interacts with xCGLs through binding to N-acetyllactosamine type N-glycans and that this interaction might make it possible to organize a lectin network involving members of different lectin families.     

Cortical spreading depression modifies components of the inflammatory cascade

As more information becomes available regarding the role of inflammation following stroke, it is apparent that some inflammatory mediators are detrimental and others are beneficial to the progression of ischemic injury. Cortical spreading depression (CSD) is known to impart some degree of ischemic tolerance to the brain and to influence the expression of many genes. Many of the genes whose expression is altered by CSD are associated with inflammation, and it appears likely that modulation of the inflammatory response to ischemia by CSD contributes to ischemic tolerance. Understanding which inflammatory processes are influenced by CSD may lead to the identification of novel targets in the effort to develop an acute treatment for stroke.