抗角蛋白自身抗体对培养角朊细胞产生白细胞介素8的影响

为探讨抗角蛋白自身抗体（AKantoAb）对角朊细胞免疫功能的影响，实验以IL-1β刺激无血清培养角朊细胞及AKautoAb作用后的角朊细胞，用夹心ELISA法检测培养上清中IL－8的产量。结果表明IL-1β可刺激角朊细胞产生IL－8，并对IL－1β有浓度依赖性，AKautoAb对角朊细胞产生IL－8有明显抑制作用。提示AKautoAb在炎性皮损角朊细胞中的沉着，可能存在有益的调节作用。     

Epidermolysis bullosa simplex Dowling-Meara due to an arginine to cysteine substitution in exon 1 of keratin 14

Epidermolysis bullosa simplex (EBS) is a blistering disorder affecting the basal layer of the epidermis usually inherited in an autosomal dominant fashion. Most cases are caused by mutations in the genes encoding keratin 5 (K5) and keratin 14 (K14) and are characterized by cytolysis within the basal layer of the epidermis. We report a patient manifesting the Dowling-Meara variant of EBS in whom we characterized a cytosine to thymine transition at codon 125 (R125C) in K14. This missense mutation is located at the amino terminus of the helical rod domain of the keratin 14 molecule, resulting in defective pairing with K5, thereby disrupting keratin tonofibril integrity.     

PCR-DNA直接测序检测1例单纯型大疱性表皮松解症Weber-Cockayne亚型(WC-EBS)患者角蛋白K5基因点突变

单纯型大疱性表皮松解症（ＥＢＳ）是一组常染色体显性的遗传性疾病，研究表明本病存在角蛋白Ｋ５／Ｋ１４基因点突变。ＥＢＳ的各个亚型突变发生部位有一定差异，其中Ｗｅｂｅｒ－Ｃｏｃｋａｙｎｅ亚型（ＷＣ－ＥＢＳ）突变多位于Ｋ５／Ｋ１４的连接区Ｌ１－２。本研究设计了扩增Ｋ５基因Ｌ１－２区ＤＮＡ片段的引物，应用ＰＣＲ对－ＷＣ－ＥＢＳ家系的患者及未发病成员进行扩增。ＰＣＲ产物测序发现患者Ｋ５第３４６密码子发生了Ａ→Ｃ的碱基替换，导致色氨酸（ＴＡＴ）变成丝氨酸（ＴＣＴ），而未发病成员则未见有碱基突变。结果表明，通过ＰＣＲ结合ＤＮＡ直接测序不失为快速、准确检测基因突变的方法。此外，连接区在角蛋白结构中不如螺旋区重要，因而此区基因突变对角蛋白二聚体形成的影响不大，这与临床上ＷＣ－ＥＢＳ病情相对较轻是一致的。     

Cloning and characterization of antikeratin human antibodies using a semisynthetic phage antibody library

Antikeratin autoantibodies (AK auto Abs) are very important elements of the human immune system. To improve the outcome of studies on AK auto Abs, it is necessary to generate antikeratin human monoclonal antibodies. The purpose of present study was to isolate antikeratin human monoclonal antibodies by panning a phage antibody library. A semisynthetic phage antibody library with capacity of 4.0×10⁸ members was previously constructed. Panning of the library was performed against human epidermal keratin extracted from psoriatic scales. At the last round of the panning, individual colonies were grown in culture for expression of phage antibodies. Their binding activities and specificities to keratin were determined by ELISA, and positive clones were analyzed by DNA fingerprinting. The selected clones were induced with IPTG to express soluble Fab fragments, which were further characterized by ELISA, immunohistochemistry and Western blotting. Finally, DNA sequencing of the variable regions was performed. A human antibody clone which was able to express soluble Fab fragments and recognize Mr 46,000 keratin (K17) was isolated. DNA sequencing demonstrated that the VH and VL of the antibody came from the human VH1 and V2 families, respectively. We conclude that phage antibody library technology is a powerful way to generate human monoclonal antibodies. The antikeratin antibody we isolated in the present study would be useful in the research on AK auto Abs.     

Spinal cord tau pathology in cervical spondylotic myelopathy

We conducted an immunohistochemical and ultrastructural examination of the spinal cords from 11 cases of cervical spondylotic myelopathy (CSM), together with those from 11 age- and sex-matched control subjects. Immunostaining with AT8 antibody revealed various numbers of tau-positive neuropil thread-like structures (NTSs), often demonstrating a conspicuous astrocytic foot-like perivascular or subpial arrangement, and glial cells with short and thick processes, so-called thorn-shaped astrocytes (TSAs), in the affected cervical cords in 8 of the 11 CSM cases (73%). A number of tau-positive neuronal cytoplasmic pretangles/tangles were also found in the gray matter in all the CSM cases (100%). No such astrocytic or neuronal tau lesions were found in the control subjects. The tau deposited in the NTSs and TSAs was predominantly 4-repeat tau, whereas the neuronal cytoplasmic pretangles/tangles contained both 3-repeat and 4-repeat tau. Ultrastructurally, paired helical filaments about 20 nm wide, together with glial filaments, were detected occasionally in the astrocytic processes. In conclusion, the present findings indicate that astrocytic and neuronal tau lesions appear in the affected cervical cord during the disease process of CSM.     

Ultrastructural Study of Symmetrical Acral Keratoderma

Abstract

Background: Symmetrical acral keratoderma is characterized by symmetrical brown hyperkeratotic patches on the acral extremities. However, no studies about its electron microscopic examination have been documented.

Objective: Our study was performed to further characterize the histopathology of symmetrical acral keratoderma.

Methods: A biopsy was taken from brown hyperkeratotic patches on the wrists. Investigative studies included light and electron microscopy.

Results: Light microscopy showed epidermal basket-weave hyperkeratosis and acanthosis. Ultrastructurally, the epidermis was thickened by acanthosis and compact stratum corneum. The horny cell layers were remarkably thicker in clinical affected skin than in adjacent clinically unaffected and healthy skin. The keratin filaments were remarkably clumped or aggregated and irregularly distributed in the horny, spinous, granular and basal cell layers. The tonofilaments formed tight clumps or aggregated at the perinuclear cytoplasm.

Conclusion: The main ultrastructural features of symmetrical acral keratoderma were epidermal hyperkeratosis and abnormalities of the keratin filaments and tonofilaments.     

正常口腔粘膜上皮的细胞角蛋白的表达

目的：探讨CK10、CK13、CK18、CK19在正常口腔粘膜中的表达。方法 ：正常颊粘膜、舌腹口底粘膜、附着眼粘膜、舌背粘膜标本各8例，10％中性福尔马林液固定，石蜡包埋，5μm连续切片，用LSAB法进行CK10、CK13、CK18、CK19的免疫组化染色。结果：正常颊粘膜和舌腹口底粘膜基底细胞层散在表达CK19，基底上层表达CK13；正常附着龈粘膜基底上层表达CK10；正常舌背粘膜基底细胞层可散在表达CK19，基底上层表达CK10和／或CK13。结论：不同部位的正常口腔粘膜表达的CK种类和形式不同。     

Three-dimensional structure of Alzheimer's neurofibrillary tangles of the aged human brain revealed by the quick-freeze,deep-etch and replica method

Summary The three-dimensional structure of Alzheimer's neurofibrillary tangles in the pyramidal cells of the hippocampus and in the nerve cells of the parahippocampal gyrus was examined by the quick-freeze, deep-etch and replica method. The tangles consisted of either parallel bundles of or randomly arranged paired helical filaments (PHF), occupying the perikaryotic cytoplasm and extending to the dendritic processes. On the stereophotographs the PHF, measuring 28 to 36 nm in width, had two component filaments of 14 to 18 nm in diameter which were coiled anti-clockwise (left-handed) around each other with periodicity of 70 to 90 nm. The PHF in compact parallel bundles were cross-linked to each other with thin filaments, of about 6 nm in diameter, at relatively regular intervals. Randomly arranged PHF had no cross-bridges or side arms. Straight-type tangles of about 24 nm in diameter were rarely found in the dendritic processes. There were no discernible differences between the PHF of the patients with senile dementia of Alzheimer type and those of nondemented brains.     

细胞角蛋白与肿瘤的关系

细胞角蛋白(CK)具有维持正常细胞形态和功能的重要作用.临床上,细胞角蛋白被作为肿瘤标记物用于多种肿瘤的诊断.近年来许多研究发现细胞角蛋白的异常表达、磷酸位点的基因突变以及过度磷酸化与肿瘤的发生机制相关,研究两者之间的关系,对于肿瘤的诊断、治疗、预后有重要的作用.     

Induction of epitopes associated with neurofibrillary tangles in clonal mouse neuroblastoma (S20Y) cells

Summary Accumulation of paired helical filaments (PHF) in neurofibrillary tangles is a key neuropathological hallmark in Alzheimer's disease (AD). To date, PHF have been found primarily in humans. Cultured murine cholinergic neuroblastoma (S20Y) cells, following exposure to a serum-free medium or a differentiation medium, developed immunoreactivity to anti-PHF antibodies, and to the Alz-50 by immunocytochemical and immunoblot analyses. Electron microscopic examination revealed abundant fascicles of 10-nm filaments coursing tortuously amongst organelles, such as mitochondria, endoplasmic reticulum and dense-core vesicles, in perikarya and in neuritic extensions. However, subcellular structures identical or similar to PHF could not be found in these non-human cells. This convenient cell culture model may prove to be useful for studying certain aspects of the mechanisms underlying the abnormal cytoskeletal alterations which are characteristic of AD and related neurodegenerative disorders.Supported by grants from the Overbrook Foundation, the Will Rogers Institute, the Dr. I. Fund Foundation, the Winifred Masterson Burke Relief Foundation, the Alzheimer's Disease Research Program of the American Health Assistance Foundation and the National Institute of Aging (AG03853)