纤维样变肾小球病：——附四例报告

报告４例纤维样变肾小球病。４例病人无系统性疾病，主要临床表现为高血压，大量蛋白尿、镜下血尿。２例有不同程度的肾功能减退。光镜下病理类型为膜增殖性肾炎（２例）、膜性肾病（１例）和系膜增生性肾炎（１例），病理改变均较重，刚果红染色阴性；免疫荧光多数病例（３１４例）以ＩｇＧ、Ｃ＿３为主，呈颗粒样在肾小球系膜区和（或）沿肾小球毛细血管壁沉积；电镜下可见大量纤维样物质在肾小球系膜区和（或）肾小球基底膜内分布，纤维样物质直径经图象分析仪测量为２０．５０±１０．３７ｎｍ。本病诊断主要依靠超微结构检查。本病为国内首次报道。     

IgA肾病的免疫电镜的研究

IgA肾病是一个以IgA沉积于肾小球系膜区为特点的疾病。至于在免疫荧光所见的免疫沉积物与电镜所见的电子致密物之间的关系及系膜增殖的程度与 IgA沉积物的量之间的关系如何,至今未见到报告。本文对16例IgA肾病进一步作了研究。光镜检查其病变分为三类,即微小病变2例,局灶性系膜增殖性肾炎3例及弥漫性系膜增殖性肾炎11例。同时进行免疫荧光、电镜及免疫电镜检查。结果发现16例电镜见到的电子致密物部位与免疫电镜所见的IgA沉积部位相一致。免疫电镜所见与免疫荧光所见的符合率也在95%以上。微小病变和局灶性系膜增殖性肾炎     

18例非典型膜性肾病的临床病理特点

目的:本文旨在探讨临床上发现的一类伴特殊电镜表现的非典型膜性肾病的临床病理特征。方法:回顾性分析解放军南京总医院肾脏科2012年1月至2016年8月间经肾活检光镜表现为肾小球病变轻,电镜下以基膜上皮侧少量或散在的规则圆形致密颗粒沉积为主,诊断为非典型膜性肾病的18例患者,观察其临床、实验室指标及病理特点。结果:(1)女性14例,男性4例,平均年龄为37. 17±15. 22岁,临床主要表现为水肿、不同程度的尿蛋白(4.38±3.78)g/24h,3例患者伴镜下血尿(18～140万/ml,多形性),无肉眼血尿,16例患者尿N-乙酰-β-D-氨基葡萄糖苷酶(NAG酶)升高,8例患者尿视黄醇结合蛋白(RBP)升高,所有患者抗磷脂酶A2受体(PLA_2R)抗体阴性;(2) 10例患者有美白化妆品使用史或染发史,余无明确诱因;(3)肾组织学特点:光镜下肾小球轻度系膜细胞增生和系膜基质增多,6例患者可见肾小管间质急性病变。免疫荧光示IgG或C3沿肾小球毛细血管袢呈颗粒样沉积,IgG亚型以IgG1沉积为主。电镜下均见肾小球基膜上皮侧少量或散在的中高密度电子致密物沉积,致密物呈类圆形或圆形,有不同程度的足突融合;(4)给予泼尼松或联合雷公藤、血管紧张素Ⅱ受体拮抗剂(ARB)等治疗,随访过程中11例患者尿检转阴。结论:临床上发现光镜表现为肾小球病变轻微,结合免疫荧光和电镜特点最后诊断为非典型膜性肾病,大多数有美白化妆品使用史及染发史,临床上预后较好。     

泌尿系统疾病

一58,中国医学文摘·内科学1997年第18卷第1期 970554 104例肾活检病理与临床分析/王文新一//临床医学一19弱,le(5),一1~3 肾活检10吕例,成功104例(9石.3%),失败4例,无1例出现严重术后并发症。病理类型共19种,诊断原发性肾小球疾病71例(68%),以系膜增殖性肾炎最多见,坛八肾病次之,膜性肾病第三,少数为膜增殖性肾小球肾炎亚型及毛细血管内增生性肾小球肾炎‘各型肾小球疾病中,临床表现以肾病综合征最常见占62.5%,其次为慢性肾炎占27%,IgA肾病及紫舞性肾炎临床表现多样。用激素治疗各型患者,微小病变型肾病近期疗效最佳,膜性肾病次之,系膜膜…     

IgA肾病的诊断与治疗探讨

IgA肾病的病变主要在系膜部位,故又称系膜性IgA肾病。其在国外沿用的病名颇多,如IgA—IgG肾病、IgA相关肾小球肾炎、Berger病、血管系膜性IgA肾炎、良性反复发作性血尿和局灶性肾炎伴系膜IgA沉积等。由于目前国内尚不能普及肾穿刺活检、电镜检查、免疫荧光染色,故本病常被误诊为其它类型的肾小球疾病。     

膜性肾病的治疗

膜性肾病(MN)是一病理学诊断名称,组织学特征是肾小球毛纲血管壁弥漫性增厚,但无系膜、内皮或上皮细胞增殖,银染色可见肾小球基底膜(GBM)上有钉突存在,免疫荧光检查可见毛细血管袢有IgG及C_3呈颗粒状沉积,常呈团状或块状分布,在系膜区也可散见。电镜下可见GBM增厚及有不连续的电子致密物。在PASM-Masson染色下可按病变发展过程分为四期。I期,基膜下有散在的红色沉积物(沉积于     

5例伴单克隆免疫球蛋白沉积的增生性肾小球肾炎临床及病理分析

目的 探讨伴单克隆免疫球蛋白沉积的增生性肾小球肾炎(PGNMID)的临床及病理特点。方法 回顾性分析5例PGNMID患者的临床特征及肾脏病理资料。结果 5例PGNMID患者中,男4例,女1例,年龄47～68岁,病程2.5～60.0个月,2例病程超过1年,5例均呈肾病综合征,伴高血压病史,4例镜下血尿,3例肾功能异常。仅1例血清免疫球蛋白游离轻链比率异常。光镜下,5例患者的肾脏均呈膜增生性肾小球肾炎病变;免疫荧光下,PGNMID患者肾小球仅见免疫球蛋白G3和κ轻链沉积;电镜显示PGNMID患者肾脏系膜、内皮下和(或)上皮下颗粒状沉积物。结论 PGNMID患者常出现肾病综合征和肾功能下降,需尽早行肾脏病理检查,以提高PGNMID的诊断率,减少漏诊和误诊。     

抗肾小球基膜肾炎合并膜性肾病

青年男性患者,临床表现急进性肾炎综合征伴大量蛋白尿、低蛋白血症,血清抗肾小球基膜(GBM)抗体阳性,肾活检组织学呈新月体肾炎合并膜性肾病,免疫病理IgG呈线状及颗粒状沉积于血管袢,电镜下见GBM上皮侧电子致密物沉积,最终诊断为抗GBM肾炎合并膜性肾病.     

成人微小病变肾病的治疗

微小病变肾病（MCD）指表现为肾病综合征、光镜下肾小球无病变（或仅轻微系膜病变）、免疫荧光阴性（或微弱的C3,IgM染色）、电镜下足突融合但无电子致密物沉积。微小病变肾病是儿童肾病综合征最常见的病因,在成人肾病综合征中占10％～15％。     

膜性肾小球肾炎的研究进展

膜性肾病或膜性肾小球肾炎是免疫复合物沉积在肾小球基底膜和脏层上皮之间,并引起基底膜弥漫性增厚的慢性原发性肾小球疾病。其临床特征为大量蛋白尿或肾病综合征。近年来,临床上对该病的研究日趋重视。现概述如下。     

Aberration-free three-dimensional multiphoton imaging of neuronal activity at kHz rates

Multiphoton microscopy is a powerful tool in neuroscience, promising to deliver important data on the spatiotemporal activity within individual neurons as well as in networks of neurons. A major limitation of current technologies is the relatively slow scan rates along the z direction compared to the kHz rates obtainable in the x and y directions. Here, we describe a custom-built microscope system based on an architecture that allows kHz scan rates over hundreds of microns in all three dimensions without introducing aberration. We further demonstrate how this high-speed 3D multiphoton imaging system can be used to study neuronal activity at millisecond resolution at the subcellular as well as the population level.     

Why inclusion bodies do assume different locations in thalassaemic erythrocytes

Summary The reported scanning (SEM), transmission (TS), and freeze-etching (FE) electron microscopic studies have agreed in confirming that in thalassaemic erythrocytes, previously incubated with brilliant cresyl blue (BCB), the unpaired alpha chains precipitate in the central portions of the cell whereas excess beta chains locate in the submembranous regions. This is due to the fact that beta chains, possessing two thiols instead of only one (as in alpha chains), are more liable to bind to similar groups contained in the inner red cell leaflet. Less soluble alpha chains tend to form inter-chain bridges and thus precipitate centrally. SEM observations have given evidence that on the surface of the affected red cells denaturated alpha chains give rise to large and shallow invaginations whereas denatured beta chains lead to a diffuse wrinkled appearance. The causes of the different SEM aspects have been suggested.     

Just Seeing Is Not Enough for Believing: Immunolabelling as Indisputable Proof of SARS-CoV-2 Virions in Infected Tissue

Background: There is increasing evidence that identification of SARS-CoV-2 virions by transmission electron microscopy could be misleading due to the similar morphology of virions and ubiquitous cell structures. This study thus aimed to establish methods for indisputable proof of the presence of SARS-CoV-2 virions in the observed tissue. Methods: We developed a variant of the correlative microscopy approach for SARS-CoV-2 protein identification using immunohistochemical labelling of SARS-CoV-2 proteins on light and electron microscopy levels. We also performed immunogold labelling of SARS-CoV-2 virions. Results: Immunohistochemistry (IHC) of SARS-CoV-2 nucleocapsid proteins and subsequent correlative microscopy undoubtedly proved the presence of SARS-CoV-2 virions in the analysed human nasopharyngeal tissue. The presence of SARS-CoV-2 virions was also confirmed by immunogold labelling for the first time. Conclusions: Immunoelectron microscopy is the most reliable method for distinguishing intracellular viral particles from normal cell structures of similar morphology and size as virions. Furthermore, we developed a variant of correlative microscopy that allows pathologists to check the results of IHC performed first on routinely used paraffin-embedded samples, followed by semithin, and finally by ultrathin sections. Both methodological approaches indisputably proved the presence of SARS-CoV-2 virions in cells.     

Correlative 3D superresolution fluorescence and electron microscopy reveal the relationship of mitochondrial nucleoids to membranes

Microscopic images of specific proteins in their cellular context yield important insights into biological processes and cellular architecture. The advent of superresolution optical microscopy techniques provides the possibility to augment EM with nanometer-resolution fluorescence microscopy to access the precise location of proteins in the context of cellular ultrastructure. Unfortunately, efforts to combine superresolution fluorescence and EM have been stymied by the divergent and incompatible sample preparation protocols of the two methods. Here, we describe a protocol that preserves both the delicate photoactivatable fluorescent protein labels essential for superresolution microscopy and the fine ultrastructural context of EM. This preparation enables direct 3D imaging in 500- to 750-nm sections with interferometric photoactivatable localization microscopy followed by scanning EM images generated by focused ion beam ablation. We use this process to "colorize" detailed EM images of the mitochondrion with the position of labeled proteins. The approach presented here has provided a new level of definition of the in vivo nature of organization of mitochondrial nucleoids, and we expect this straightforward method to be applicable to many other biological questions that can be answered by direct imaging.     

Charge transport and intrinsic fluorescence in amyloid-like fibrils

The self-assembly of polypeptides into stable, conductive, and intrinsically fluorescent biomolecular nanowires is reported. We have studied the morphology and electrical conduction of fibrils made of an elastin-related polypeptide, poly(ValGlyGlyLeuGly). These amyloid-like nanofibrils, with a diameter ranging from 20 to 250 nm, result from self-assembly in aqueous solution at neutral pH. Their morphological properties and conductivity have been investigated by atomic force microscopy, scanning tunneling microscopy, and two-terminal transport experiments at the micro- and nanoscales. We demonstrate that the nanofibrils can sustain significant electrical conduction in the solid state at ambient conditions and have remarkable stability. We also show intrinsic blue-green fluorescence of the nanofibrils by confocal microscopy analyses. These results indicate that direct (label-free) excitation can be used to investigate the aggregation state or the polymorphism of amyloid-like fibrils (and possibly of other proteinaceous material) and open up interesting perspectives for the use of peptide-based nanowire structures, with tunable physical and chemical properties, for a wide range of nanobiotechnological and bioelectronic applications.     

Application of Super-Resolution and Advanced Quantitative Microscopy to the Spatio-Temporal Analysis of Influenza Virus Replication

With an estimated three to five million human cases annually and the potential to infect domestic and wild animal populations, influenza viruses are one of the greatest health and economic burdens to our society, and pose an ongoing threat of large-scale pandemics. Despite our knowledge of many important aspects of influenza virus biology, there is still much to learn about how influenza viruses replicate in infected cells, for instance, how they use entry receptors or exploit host cell trafficking pathways. These gaps in our knowledge are due, in part, to the difficulty of directly observing viruses in living cells. In recent years, advances in light microscopy, including super-resolution microscopy and single-molecule imaging, have enabled many viral replication steps to be visualised dynamically in living cells. In particular, the ability to track single virions and their components, in real time, now allows specific pathways to be interrogated, providing new insights to various aspects of the virus-host cell interaction. In this review, we discuss how state-of-the-art imaging technologies, notably quantitative live-cell and super-resolution microscopy, are providing new nanoscale and molecular insights into influenza virus replication and revealing new opportunities for developing antiviral strategies.     

Single molecule fluorescence and force microscopy

The investigation of biomolecules has entered a new age since the development of methodologies capable of studies at the level of single molecules. In biology, most molecules show a complex dynamical behavior, with individual motions and transitions between different states occurring highly correlated in space and time within an arrangement of various elements. Recent advances in the development of new microscopy techniques with sensitivity at the single molecule have gained access to essentially new types of information obtainable from imaging biomolecular samples. These methodologies are described here in terms of their applicability to the in vivo detection and visualization of molecular processes on surfaces, membranes, and cells. First examples of single molecule microscopy on cell membranes revealed new basic insight into the lateral organization of the plasma membrane, providing the captivating perspective of an ultra-sensitive methodology as a general tool to study local processes and heterogeneities in living cells.     

Pineal Gland Morphology in Rats With Experimentally Induced Protein-Calorie Malnutrition

The morphology of the pineal gland was studied in protein-calorie-malnourished (PCM) rats. Twenty-day-old male Sprague-Dawley rats were placed in a 14:10 photoperiod and fed either an 8% low protein diet (LPD) or a standard laboratory diet (SLD) containing 27% protein for 30 d. At 50 d of age, rats from both animal groups were sacrificed at 0900 h and at 2400 h, and the pineal glands were immersion-fixed for either light or electron microscopic analysis. The cytoplasm and nuclei of the pinealocytes from the SLD-fed rats were consistently larger than those of the animals maintained on the LPD. Additionally, the lipid droplets were larger and more prominent in the controls at both 0900 h and 2400 h. Even though the size of these inclusions did not vary among animals given the same diet as a function of the time of sacrifice, they were more numerous in both the well-fed and malnourished rats during the dark phase of the photoperiod. In contrast neither diet nor sampling time affected the size or number of pinealocyte mitochondria. These morphological observations lend further support to the premise than PCM impairs the cellular activity of the pinealocytes.     

High-resolution restoration of 3D structures from widefield images with extreme low signal-to-noise-ratio

Four-dimensional fluorescence microscopy—which records 3D image information as a function of time—provides an unbiased way of tracking dynamic behavior of subcellular components in living samples and capturing key events in complex macromolecular processes. Unfortunately, the combination of phototoxicity and photobleaching can severely limit the density or duration of sampling, thereby limiting the biological information that can be obtained. Although widefield microscopy provides a very light-efficient way of imaging, obtaining high-quality reconstructions requires deconvolution to remove optical aberrations. Unfortunately, most deconvolution methods perform very poorly at low signal-to-noise ratios, thereby requiring moderate photon doses to obtain acceptable resolution. We present a unique deconvolution method that combines an entropy-based regularization function with kernels that can exploit general spatial characteristics of the fluorescence image to push the required dose to extreme low levels, resulting in an enabling technology for high-resolution in vivo biological imaging.The study of dynamic processes is an important facet of cell biology research. Fluorescently tagged proteins combined with four-dimensional fluorescence microscopy, which records 3D image information as a function of time, provide a powerful framework for studying the dynamics of molecular processes in vivo. One of the most crucial challenges in 4D fluorescence microscopy is to ensure that normal biological function is not significantly perturbed as a result of the high doses of illumination (phototoxicity) incurred during 4D imaging. Recent work indicates that the maximal photon dose that avoids biological perturbation is 100- to 1,000-fold lower than that typically used for in vivo imaging (1). Dose limitations are even more challenging, given the desire to densely sample in time or to record over extended periods, especially in the context of analyzing multiple subcellular components via multiwavelength imaging.Under normal imaging conditions, widefield microscopy combined with image restoration using deconvolution methods provides an excellent modality for multiwavelength 4D imaging as it makes very efficient use of the illuminating photons. However, its effectiveness, in particular its ability to resolve subcellular detail sufficiently in the presence of noise, is limited by the performance of the deconvolution method. Such limitations can seriously degrade image quality at the low signal levels required for unperturbed in vivo imaging. The noise behavior of the deconvolution algorithm is determined by the efficiency of the noise stabilization term, known as the regularization functional. In particular, the functional’s ability to discriminate the noise-related high frequencies from weak high frequencies in the signal ultimately determines the final resolution of the deconvolution. Currently used noise-stabilization techniques are largely based on ad hoc formulations and perform poorly, leading to a serious loss of resolution at the low signal-to-noise ratios required to maintain the illumination at safe levels during multiwavelength 4D imaging. Surmounting this problem would dramatically increase the amount of biological data that could be safely acquired, paving the way for a much deeper understanding of the dynamics of biological processes.We propose a unique deconvolution method that uses a regularization functional constructed using an entropy-based formalism that is tailored to exploit general spatial characteristics of the fluorescence images combined with the more robust use of second-order derivatives in the regularization functional. This entropic-based regularization suppresses large amounts of noise while at the same time preserving the essential details. Hence the method brings out details that are nearly invisible in the raw extremely noisy images and yields a substantially improved resolution. Using several datasets of fixed samples recorded at high and low doses, we quantitatively study the performance of our method, using Fourier shell correlation methods, and demonstrate that entropy-regularized deconvolution (ER-Decon) reveals considerably more detail of the underlying structure compared with existing methods.     

大肠腺管开口分型的超微结构研究

目的通过对大肠息肉的超微结构观察,分析其表面腺管开口形态（pit pattern）的演化过程与息肉性质发生改变的关系。方法对50例大肠息肉患者行结肠放大内镜检查,结合内镜下黏膜染色对病灶行pit pattern分型,内镜或手术切除病灶,所有样本分别行组织病理学分析、扫描及透射电镜观察。结果本组50例样本,pit pattern记录Ⅰ型10个、Ⅱ型8个、Ⅲ型15个、Ⅳ型9个、Ⅴ型5个、混合型3个。组织学诊断验证其阳性预测准确率达86％（43／50）,鉴别腺瘤性息肉的准确率达94％（30／32）。扫描电镜下Ⅰ、Ⅱ型隐窝规则洞状,隐窝周围细胞大小一致,吸收细胞微绒毛多,杯状细胞丰富;Ⅲ、Ⅳ、Ⅴ型隐窝逐渐变形,表面细胞及组织异型程度不断加重。透射电镜下Ⅰ、Ⅱ型腺上皮细胞排列整齐,连接紧密,腔面微绒毛规则,胞浆线粒体、内质网丰富,核卵圆形位于基底部,基底膜平滑连续;Ⅲ、Ⅳ、Ⅴ型腺上皮细胞绒毛逐渐稀疏紊乱,胞浆线粒体肿胀,内质网扩张,溶酶体数量增加,核异型、核分裂明显加重,基板逐渐出现中断或消失。结论pit pattern的变化受大肠隐窝表面细胞的形态、比例以及构成等因素的影响,腺上皮细胞的超微结构变化是pit pattern形态和息肉性质发生改变的物质基础。