Journal of Clinical Laboratory Analysis最新文献_第10页

Dynein Light Intermediate Chains Exhibit Different Arginine Methylation Patterns Dynein轻中间链表现出不同的精氨酸甲基化模式

IF 2.7 4区医学 Q2 MEDICAL LABORATORY TECHNOLOGY

Journal of Clinical Laboratory Analysis

Pub Date : 2024-03-25 DOI: 10.1002/jcla.25030

Weiwen Bu, Jie Di, Junkui Zhao, Ruming Liu, Yue Wu, Jie Ran, Te Li

Background

The motor protein dynein is integral to retrograde transport along microtubules and interacts with numerous cargoes through the recruitment of cargo-specific adaptor proteins. This interaction is mediated by dynein light intermediate chain subunits LIC1 (DYNC1LI1) and LIC2 (DYNC1LI2), which govern the adaptor binding and are present in distinct dynein complexes with overlapping and unique functions.

Methods

Using bioinformatics, we analyzed the C-terminal domains (CTDs) of LIC1 and LIC2, revealing similar structural features but diverse post-translational modifications (PTMs). The methylation status of LIC2 and the proteins involved in this modification were examined through immunoprecipitation and immunoblotting analyses. The specific methylation sites on LIC2 were identified through a site-directed mutagenesis analysis, contributing to a deeper understanding of the regulatory mechanisms of the dynein complex.

Results

We found that LIC2 is specifically methylated at the arginine 397 residue, a reaction that is catalyzed by protein arginine methyltransferase 1 (PRMT1).

Conclusions

The distinct PTMs of the LIC subunits offer a versatile mechanism for dynein to transport diverse cargoes efficiently. Understanding how these PTMs influence the functions of LIC2, and how they differ from LIC1, is crucial for elucidating the role of dynein-related transport pathways in a range of diseases. The discovery of the arginine 397 methylation site on LIC2 enhances our insight into the regulatory PTMs of dynein functions.

背景：运动蛋白动力蛋白（dynein）是沿微管逆向运输不可或缺的部分，它通过招募货物特异性适配蛋白与许多货物相互作用。这种相互作用由动力蛋白轻中间链亚基 LIC1（DYNC1LI1）和 LIC2（DYNC1LI2）介导，它们控制适配体的结合，存在于不同的动力蛋白复合物中，具有重叠和独特的功能：我们利用生物信息学方法分析了 LIC1 和 LIC2 的 C 端结构域（CTD），发现它们具有相似的结构特征，但翻译后修饰（PTM）却各不相同。通过免疫沉淀和免疫印迹分析研究了LIC2的甲基化状态以及参与这种修饰的蛋白质。通过定点突变分析确定了 LIC2 上的特定甲基化位点，有助于加深对动力蛋白复合体调控机制的理解：结果：我们发现 LIC2 在精氨酸 397 残基上被特异性甲基化，该反应由蛋白精氨酸甲基转移酶 1（PRMT1）催化：结论：LIC 亚基的不同 PTM 为动力蛋白高效运输各种货物提供了一种多功能机制。了解这些 PTM 如何影响 LIC2 的功能，以及它们与 LIC1 的区别，对于阐明与动力蛋白相关的转运途径在一系列疾病中的作用至关重要。LIC2 上精氨酸 397 甲基化位点的发现增强了我们对动力蛋白功能调控 PTM 的洞察力。

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引用次数: 0

INPP5E Regulates the Distribution of Phospholipids on Cilia in RPE1 Cells INPP5E 调节 RPE1 细胞纤毛上磷脂的分布。

IF 2.7 4区医学 Q2 MEDICAL LABORATORY TECHNOLOGY

Journal of Clinical Laboratory Analysis

Pub Date : 2024-03-21 DOI: 10.1002/jcla.25031

Denghui Zhai, Lamei Li, Cheng Chen, Xue Wang, Ruming Liu, Ying Shan

Background

Primary cilia are static microtubule-based structures protruding from the cell surface and present on most vertebrate cells. The appropriate localization of phospholipids is essential for cilia formation and stability. INPP5E is a cilia-localized inositol 5-phosphatase; its deletion alters the phosphoinositide composition in the ciliary membrane, disrupting ciliary function.

Methods

The EGFP-2xP4M^SidM, PH^PLCδ1-EGFP, and SMO-tRFP plasmids were constructed by the Gateway system to establish a stable RPE1 cell line. The INPP5E KO RPE1 cell line was constructed with the CRISPR/Cas9 system. The localization of INPP5E and the distribution of PI(4,5)P₂ and PI4P were examined by immunofluorescence microscopy. The fluorescence intensity co-localized with cilia was quantified by ImageJ.

Results

In RPE1 cells, PI4P is localized at the ciliary membrane, whereas PI(4,5)P₂ is localized at the base of cilia. Knocking down or knocking out INPP5E alters this distribution, resulting in the distribution of PI(4,5)P₂ along the ciliary membrane and the disappearance of PI4P from the cilia. Meanwhile, PI(4,5)P₂ is located in the ciliary membrane labeled by SMO-tRFP.

Conclusions

INPP5E regulates the distribution of phosphoinositide on cilia. PI(4,5)P₂ localizes at the ciliary membrane labeled with SMO-tRFP, indicating that ciliary pocket membrane contains PI(4,5)P₂, and phosphoinositide composition in early membrane structures may differ from that in mature ciliary membrane.

背景：初级纤毛是从细胞表面伸出的基于微管的静态结构，存在于大多数脊椎动物细胞中。磷脂的适当定位对纤毛的形成和稳定性至关重要。INPP5E 是一种纤毛定位的肌醇 5-磷酸酶；它的缺失会改变纤毛膜中的磷脂组成，从而破坏纤毛的功能：方法：通过Gateway系统构建了EGFP-2xP4MSidM、PHPLCδ1-EGFP和SMO-tRFP质粒，以建立稳定的RPE1细胞系。利用 CRISPR/Cas9 系统构建了 INPP5E KO RPE1 细胞系。免疫荧光显微镜检测了 INPP5E 的定位以及 PI(4,5)P2 和 PI4P 的分布。用 ImageJ 对与纤毛共定位的荧光强度进行量化：结果：在 RPE1 细胞中，PI4P 定位于纤毛膜，而 PI(4,5)P2 定位于纤毛基部。敲低或敲除 INPP5E 会改变这种分布，导致 PI(4,5)P2 沿纤毛膜分布，PI4P 从纤毛中消失。同时，PI(4,5)P2位于SMO-tRFP标记的纤毛膜上：结论：INPP5E调节磷酸肌醇在纤毛上的分布。PI(4,5)P2定位于SMO-tRFP标记的纤毛膜上，这表明纤毛袋膜含有PI(4,5)P2，早期膜结构中的磷酸肌醇组成可能与成熟纤毛膜中的磷酸肌醇组成不同。

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引用次数: 0

Molecular Characterization of α- and β-Thalassemia Among Children Less Than 18 Years Old in Guizhou, China 中国贵州 18 岁以下儿童α-和β-地中海贫血的分子特征。

IF 2.7 4区医学 Q2 MEDICAL LABORATORY TECHNOLOGY

Journal of Clinical Laboratory Analysis

Pub Date : 2024-03-20 DOI: 10.1002/jcla.25022

Yan Li, Jiao Jin, Yuanyuan Tuo, Pei Huang, Jing Huang, Honglan Yang, Zhixu He

Background

Thalassemia is an inherited hemolytic disease, the complications and sequelae of which have posed a huge impact on both patients and society. But limited studies have investigated the molecular characterization of α- and β-thalassemia in children from Guizhou, China.

Methods

Between January 2019 and December 2022, a total of 3301 children, aged 6 months to 18 years, suspected of having thalassemia underwent molecular analysis.

Results

Out of the total sample, 824 (25%) children were found to carry thalassemia mutations. The carrier rates of α-thalassemia, β-thalassemia, and α + β-thalassemia were determined as 8.1%, 15.6%, and 1.3%, respectively. Approximately 96.5% of the α-thalassemia gene mutations were --SEA (51%), αα^CS (20.9%), -α^3.7 (19.6%), and -α^4.2 (5.0%). The most prevalent mutations of β-thalassemia were β^CD17(A>T) (41.5%), β^{CD41-42(-TTCT)} (37.7%), and β^{IVS-II-654(C>T)} (11.3%). Additionally, we identified rare cases, including one case with αα^{Hb Nunobiki}/αα, two cases with triplicated α-thalassemia (one case with ααα/ααα and β^CD41-42/β^N and the other with ααα^-3.7/αα and βE ^CD26/β^N), and also one case with α ^Q-Thailandα/-α^4.2 and β^CD41-42/β^N.

Conclusions

Our study findings provide important insights into the heterogeneity of thalassemia carrier rates and molecular profiles among children in the Guizhou region. The findings support the development of prevention strategies to reduce the incidence of severe thalassemia in the future.

背景：地中海贫血是一种遗传性溶血性疾病，其并发症和后遗症对患者和社会都造成了巨大影响。但对中国贵州儿童α-和β-地中海贫血分子特征的研究有限：方法：2019年1月至2022年12月，对3301名6个月至18岁疑似地中海贫血患儿进行分子分析：结果：在所有样本中，发现824名（25%）儿童携带地中海贫血基因突变。α地中海贫血、β地中海贫血和α+β地中海贫血的携带率分别为 8.1%、15.6% 和 1.3%。约96.5%的α地中海贫血基因突变为--SEA（51%）、ααCS（20.9%）、-α3.7（19.6%）和-α4.2（5.0%）。β地中海贫血最常见的突变是βCD17(A>T)（41.5%）、βCD41-42(-TTCT)（37.7%）和βIVS-II-654(C>T)（11.3%）。此外，我们还发现了一些罕见病例，包括一例ααHb Nunobiki /αα，两例三重α地中海贫血（一例为ααα/ααα和βCD41-42 /βN，另一例为ααα-3.7 /αα和βE CD26 /βN），以及一例α Q-Thailand α/-α4.2和βCD41-42 /βN：我们的研究结果为了解贵州地区儿童地中海贫血携带率和分子特征的异质性提供了重要依据。研究结果有助于制定预防策略，降低重型地中海贫血的发病率。

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引用次数: 0

Association of the ESR1 (rs9340799), OLR1 (rs3736234), LIPC (rs2070895), VDR (rs2228570), and CETP (rs708272) Polymorphisms With Risk of Coronary Artery Disease in Iranian Patients 伊朗患者的 ESR1 (rs9340799)、OLR1 (rs3736234)、LIPC (rs2070895)、VDR (rs2228570) 和 CETP (rs708272) 多态性与冠状动脉疾病风险的关系。

IF 2.7 4区医学 Q2 MEDICAL LABORATORY TECHNOLOGY

Journal of Clinical Laboratory Analysis

Pub Date : 2024-03-20 DOI: 10.1002/jcla.25026

Zahra Miri Karam, Abolfazl Yari, Atefeh Najmadini, Nima Norouzi Khorasani, Rezvan Attari, Saeideh Jafarinejad-Farsangi, Mohammad Ali Miri Karam, Hamid Najafipour, Kolsoum Saeidi

Background

Coronary artery disease (CAD) is a devastating illness and a leading cause of death worldwide, primarily caused by atherosclerosis resulting from a genetic-environmental interaction. This study aimed to investigate the relationship between the ESR1 (rs9340799), OLR1 (rs3736234), LIPC (rs2070895), VDR (rs2228570), and CETP (rs708272) polymorphisms, lipid profile parameters, and CAD risk in a southeast Iranian population.

Methods

A total of 400 subjects (200 CAD patients with hyperlipidemia and 200 healthy controls) were enrolled in this case–control study. Five selected polymorphisms were genotyped using the polymerase chain reaction–restriction fragment length polymorphism (PCR-RFLP) technique.

Results

For all single nucleotide polymorphisms (SNPs), the population under study was in the Hardy–Weinberg equilibrium. The T-risk allele frequency of rs2228570 was associated with an increased risk of CAD. The TT and CT genotypes of rs2228570 had also been associated with the risk of CAD. Additionally, the TT genotype was associated with higher serum low-density lipoprotein cholesterol (LDL-c) and high-density lipoprotein cholesterol (HDL-c) levels. The GG genotype of the rs3736234 was associated with higher body mass index (BMI) and triglyceride (TG) levels, and the AA genotype of the rs708272 was associated with higher HDL-c levels. Based on these findings, we propose that the VDR (rs2228570) polymorphism was associated with serum HDL-c and LDL-c levels and may serve as potential risk factors for CAD within the Iranian population. Moreover, rs3736234 and rs708272 influence the concentrations of TG and HDL-c, respectively.

Conclusion

These findings provided insights into the complex interplay between genetic variations, cardiovascular risk, and lipid metabolism.

背景：冠状动脉疾病（CAD）是一种破坏性疾病，也是全球死亡的主要原因，其主要原因是遗传与环境相互作用导致的动脉粥样硬化。本研究旨在调查伊朗东南部人群中 ESR1 (rs9340799)、OLR1 (rs3736234)、LIPC (rs2070895)、VDR (rs2228570) 和 CETP (rs708272) 多态性、血脂谱参数和 CAD 风险之间的关系：这项病例对照研究共招募了 400 名受试者（200 名患有高脂血症的 CAD 患者和 200 名健康对照者）。采用聚合酶链式反应-限制性片段长度多态性（PCR-RFLP）技术对所选的五个多态性进行了基因分型：在所有单核苷酸多态性（SNPs）中，研究人群均处于哈代-温伯格平衡状态。rs2228570的T风险等位基因频率与CAD风险增加有关。rs2228570 的 TT 和 CT 基因型也与患 CAD 的风险有关。此外，TT 基因型与较高的血清低密度脂蛋白胆固醇（LDL-c）和高密度脂蛋白胆固醇（HDL-c）水平有关。rs3736234 的 GG 基因型与较高的体重指数（BMI）和甘油三酯（TG）水平相关，而 rs708272 的 AA 基因型与较高的 HDL-c 水平相关。基于这些发现，我们认为 VDR（rs2228570）多态性与血清高密度脂蛋白胆固醇（HDL-c）和低密度脂蛋白胆固醇（LDL-c）水平有关，并可能成为伊朗人群患 CAD 的潜在风险因素。此外，rs3736234 和 rs708272 分别影响 TG 和 HDL-c 的浓度：这些研究结果有助于深入了解遗传变异、心血管风险和脂质代谢之间复杂的相互作用。

{"title":"Association of the ESR1 (rs9340799), OLR1 (rs3736234), LIPC (rs2070895), VDR (rs2228570), and CETP (rs708272) Polymorphisms With Risk of Coronary Artery Disease in Iranian Patients","authors":"Zahra Miri Karam, Abolfazl Yari, Atefeh Najmadini, Nima Norouzi Khorasani, Rezvan Attari, Saeideh Jafarinejad-Farsangi, Mohammad Ali Miri Karam, Hamid Najafipour, Kolsoum Saeidi","doi":"10.1002/jcla.25026","DOIUrl":"10.1002/jcla.25026","url":null,"abstract":"<div>\u0000 \u0000 \u0000 <section>\u0000 \u0000 <h3> Background</h3>\u0000 \u0000 <p>Coronary artery disease (CAD) is a devastating illness and a leading cause of death worldwide, primarily caused by atherosclerosis resulting from a genetic-environmental interaction. This study aimed to investigate the relationship between the <i>ESR</i>1 (rs9340799), <i>OLR1</i> (rs3736234), <i>LIPC</i> (rs2070895), <i>VDR</i> (rs2228570), and <i>CETP</i> (rs708272) polymorphisms, lipid profile parameters, and CAD risk in a southeast Iranian population.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Methods</h3>\u0000 \u0000 <p>A total of 400 subjects (200 CAD patients with hyperlipidemia and 200 healthy controls) were enrolled in this case–control study. Five selected polymorphisms were genotyped using the polymerase chain reaction–restriction fragment length polymorphism (PCR-RFLP) technique.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Results</h3>\u0000 \u0000 <p>For all single nucleotide polymorphisms (SNPs), the population under study was in the Hardy–Weinberg equilibrium. The T-risk allele frequency of rs2228570 was associated with an increased risk of CAD. The TT and CT genotypes of rs2228570 had also been associated with the risk of CAD. Additionally, the TT genotype was associated with higher serum low-density lipoprotein cholesterol (LDL-c) and high-density lipoprotein cholesterol (HDL-c) levels. The GG genotype of the rs3736234 was associated with higher body mass index (BMI) and triglyceride (TG) levels, and the AA genotype of the rs708272 was associated with higher HDL-c levels. Based on these findings, we propose that the <i>VDR</i> (rs2228570) polymorphism was associated with serum HDL-c and LDL-c levels and may serve as potential risk factors for CAD within the Iranian population. Moreover, rs3736234 and rs708272 influence the concentrations of TG and HDL-c, respectively.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Conclusion</h3>\u0000 \u0000 <p>These findings provided insights into the complex interplay between genetic variations, cardiovascular risk, and lipid metabolism.</p>\u0000 </section>\u0000 </div>","PeriodicalId":15509,"journal":{"name":"Journal of Clinical Laboratory Analysis","volume":"38 6","pages":""},"PeriodicalIF":2.7,"publicationDate":"2024-03-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jcla.25026","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140174919","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Assessment of Bone Marrow Involvement in B-Cell non-Hodgkin Lymphoma Using Immunoglobulin Gene Rearrangement Analysis with Next-Generation Sequencing 利用新一代测序的免疫球蛋白基因重排分析评估 B 细胞非霍奇金淋巴瘤的骨髓受累情况

IF 2.7 4区医学 Q2 MEDICAL LABORATORY TECHNOLOGY

Journal of Clinical Laboratory Analysis

Pub Date : 2024-03-20 DOI: 10.1002/jcla.25027

Min Ji Jeon, Eun Sang Yu, Dae Sik Kim, Chul Won Choi, Ha Nui Kim, Jung Ah Kwon, Soo-Young Yoon, Jung Yoon

Background

Assessment of bone marrow involvement (BMI) in non-Hodgkin lymphoma (NHL) is crucial for determining patient prognosis and treatment strategy. We assessed the prognostic value of next-generation sequencing (NGS)–based immunoglobulin (Ig) gene clonality analysis as an ancillary test for BMI evaluation in NHL.

Methods

A retrospective cohort of 124 patients newly diagnosed with B-cell NHL between 2019 and 2022 was included. NGS-based Ig clonality analysis was conducted using LymphoTrak IGH FR1 Assay and IGK Assay (Invivoscribe Technologies, San Diego, CA, USA) on BM aspirate samples, and the results were compared with those of histopathological BMI (hBMI).

Results

Among the 124 patients, hBMI was detected in 16.9% (n = 21). The overall agreement of BMI between Ig clonality analyses and histopathological analysis for IGH, IGK, and either IGH or IGK was 86.3%, 92.7%, and 90.3%. The highest positive percent agreement was observed with clonal rearrangements of either IGH or IGK gene (90.5%), while the highest negative percent agreement was observed with clonal rearrangement of IGK gene (96.1%). For the prediction of hBMI, positive prediction value ranged between 59.1% and 80.0% and the negative prediction value ranged between 91.3% and 97.9%.

Conclusion

NGS-based clonality analysis is an analytic platform with a substantial overall agreement with histopathological analysis. Assessment of both IGH and IGK genes for the clonal rearrangement analysis could be considered for the optimal diagnostic performance of BMI detection in B-cell NHL.

背景：评估非霍奇金淋巴瘤（NHL）的骨髓受累情况（BMI）对于确定患者预后和治疗策略至关重要。我们评估了基于新一代测序（NGS）的免疫球蛋白（Ig）基因克隆性分析作为NHL骨髓受累评估辅助检测的预后价值：方法：纳入2019年至2022年间新诊断为B细胞NHL的124名患者的回顾性队列。使用 LymphoTrak IGH FR1 Assay 和 IGK Assay（Invivoscribe Technologies，San Diego，CA，USA）对骨髓穿刺样本进行基于 NGS 的 Ig 克隆性分析，并将结果与组织病理学 BMI（hBMI）进行比较：结果：在 124 例患者中，16.9%（21 例）检测出 hBMI。Ig克隆分析与组织病理学分析在IGH、IGK以及IGH或IGK的BMI方面的总体一致性分别为86.3%、92.7%和90.3%。IGH 或 IGK 基因克隆重排的正向一致性百分比最高（90.5%），而 IGK 基因克隆重排的负向一致性百分比最高（96.1%）。在预测 hBMI 方面，阳性预测值介于 59.1% 与 80.0% 之间，阴性预测值介于 91.3% 与 97.9% 之间：结论：基于 NGS 的克隆性分析是一种分析平台，与组织病理学分析的总体一致性很高。为了优化 BMI 检测在 B 细胞 NHL 中的诊断性能，可以考虑对 IGH 和 IGK 基因进行克隆重排分析评估。

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引用次数: 0

Dissociation Phenomenon of Erythrocyte Agglutination and Its Application to Assay of Functional Activity of the Complement System in Clinical Laboratory 红细胞凝集的解离现象及其在临床实验室检测补体系统功能活性中的应用。

IF 2.7 4区医学 Q2 MEDICAL LABORATORY TECHNOLOGY

Journal of Clinical Laboratory Analysis

Pub Date : 2024-03-20 DOI: 10.1002/jcla.25028

Xuewei Ding, Lina Liu, Guang Yang, Hui Liu

Objective

The objective of the study was to validate the dissociation phenomenon of erythrocyte agglutination which is based on erythrocyte fragments and to apply it in the functional activity assay of the complement system.

Methods

The dissociation–agglutination effect of erythrocyte fragments was validated by detecting the number of free erythrocytes after the action of erythrocyte fragments on agglutinated erythrocytes. The number of free erythrocytes produced after hemolysis of agglutinated erythrocytes caused by complements and complement activators(CAs) was detected by auto hematology analyzer and the results were indicated by mean hemoglobin concentration of erythrocytes (MCHC). We optimized the test conditions and validated the inter-batch stability, explored the resolution of the assay method, and assayed for the total complement activity (AC) and the CAs activated complement activity (ACA) in serum from patients and healthy individual groups.

Results

Erythrocyte fragments have a dissociative effect on agglutinated erythrocytes. The auto hematology analyzer was able to detect AC and ACA, where AC showed an inverse correlation with MCHC, and ACA demonstrated a positive correlation with MCHC. The inter-batch CV of AC, ACA, and ACA/AC was found to be 5%, 9%, and 11.7%, respectively, with good stability. The study found that serum samples from acute phase reaction patients showed significant differences in ACA compared with healthy individuals, with a p value of 0.018; serum samples from patients with nephrotic syndrome showed significant differences in AC, ACA, and ACA/AC compared with healthy individuals, with p values of 0.014, 0.002, and 0.041, respectively.

Conclusion

Erythrocyte fragments have dissociation–agglutination effect. The complement system immunological functional detection method, based on this effect, has potential clinical application value due to its sensitivity and accuracy.

研究目的研究目的是验证基于红细胞片段的红细胞凝集解离现象，并将其应用于补体系统的功能活性测定：方法：通过检测红细胞碎片作用于凝集红细胞后游离红细胞的数量来验证红细胞碎片的解离-凝集效应。用自动血液分析仪检测补体和补体激活剂（CAs）对凝集红细胞溶血后产生的游离红细胞数量，结果用红细胞平均血红蛋白浓度（MCHC）表示。我们优化了检测条件，验证了批间稳定性，探索了检测方法的分辨率，并检测了患者和健康人血清中的总补体活性（AC）和CAs激活的补体活性（ACA）：结果：红细胞碎片对凝集的红细胞有解离作用。自动血液分析仪能检测出 AC 和 ACA，其中 AC 与 MCHC 呈反相关，而 ACA 与 MCHC 呈正相关。研究发现，AC、ACA 和 ACA/AC 的批间 CV 分别为 5%、9% 和 11.7%，稳定性良好。研究发现，急性期反应患者血清样本的 ACA 与健康人相比有显著差异，P 值为 0.018；肾病综合征患者血清样本的 AC、ACA 和 ACA/AC 与健康人相比有显著差异，P 值分别为 0.014、0.002 和 0.041：结论：红细胞碎片具有解离-凝集作用。结论：红细胞碎片具有解离-凝集效应，基于该效应的补体系统免疫功能检测方法因其灵敏性和准确性而具有潜在的临床应用价值。

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引用次数: 0

Evaluation of Automated Magnetic Bead–Based DNA Extraction for Detection of Short Tandem Repeat Expansions With Nanopore Sequencing 评估利用纳米孔测序检测短串联重复序列扩增的基于磁珠的自动 DNA 提取。

IF 2.7 4区医学 Q2 MEDICAL LABORATORY TECHNOLOGY

Journal of Clinical Laboratory Analysis

Pub Date : 2024-03-20 DOI: 10.1002/jcla.25029

Helene Faust, Patricia Duffek, Julia Hentschel, Denny Popp

Background

Long-read technologies such as nanopore sequencing provide new opportunities to detect short tandem repeat expansions. Therefore, a DNA extraction method is necessary that minimizes DNA fragmentation and hence allows the identification of large repeat expansions. In this study, an automated magnetic bead–based DNA extraction method and the required EDTA blood storage conditions as well as DNA and sequencing quality were evaluated for their suitability for repeat expansion detection with nanopore sequencing.

Methods

DNA was extracted from EDTA blood, and subsequently, its concentration, purity, and integrity were assessed. DNA was then subjected to nanopore sequencing, and quality metrics of the obtained sequencing data were evaluated.

Results

DNA extracted from fresh EDTA blood as well as from cooled or frozen EDTA blood revealed high DNA integrity whereas storage at room temperature over 7 days had detrimental effects. After nanopore sequencing, the read length N50 values of approximately 9 kb were obtained, and based on adaptive sampling of samples with a known repeat expansion, repeat expansions up to 10 kb could be detected.

Conclusion

The automated magnetic bead–based DNA extraction was sufficient to detect short tandem repeat expansions, omitting the need for high-molecular-weight DNA extraction methods. Therefore, DNA should be extracted either from fresh blood or from blood stored in cooled or frozen conditions. Consequently, this study may help other laboratories to evaluate their DNA extraction method regarding the suitability for detecting repeat expansions with nanopore sequencing.

背景：纳米孔测序等长读技术为检测短串联重复扩增提供了新的机会。因此，有必要采用一种 DNA 提取方法，以最大限度地减少 DNA 断裂，从而鉴定大重复扩增。本研究评估了一种基于磁珠的自动 DNA 提取方法和所需的 EDTA 血液储存条件以及 DNA 和测序质量，以确定其是否适合用纳米孔测序检测重复扩增：从 EDTA 血液中提取 DNA，然后评估其浓度、纯度和完整性。然后对 DNA 进行纳米孔测序，并评估测序数据的质量指标：结果：从新鲜的乙二胺四乙酸（EDTA）血液以及冷却或冷冻的乙二胺四乙酸（EDTA）血液中提取的 DNA 显示出较高的 DNA 完整性，而在室温下保存 7 天以上则会产生不利影响。纳米孔测序后，读长 N50 值约为 9 kb，根据对已知重复扩增样本的自适应取样，可检测到重复扩增达 10 kb：结论：基于磁珠的自动 DNA 提取足以检测短串联重复扩增，无需使用高分子量 DNA 提取方法。因此，DNA 应从新鲜血液或在冷却或冷冻条件下储存的血液中提取。因此，本研究可帮助其他实验室评估其DNA提取方法是否适合用纳米孔测序法检测重复扩增。

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引用次数: 0

Establishment and methodological evaluation of a chemiluminescence assay for detection of anti-envelope protein (E1, E2) antibodies in the serum of hepatitis C virus-infected patients 建立和评估用于检测丙型肝炎病毒感染者血清中抗包膜蛋白（E1、E2）抗体的化学发光测定法

IF 2.7 4区医学 Q2 MEDICAL LABORATORY TECHNOLOGY

Journal of Clinical Laboratory Analysis

Pub Date : 2024-03-15 DOI: 10.1002/jcla.25011

Ningning Wang, Qingqing Liu, Feihu Che, Qingyang Sun, Yue Wang, Chunli Yang, Yuzhu Dai, Jun Cheng

Background

To establish a chemiluminescence method for detecting anti-E1 and anti-E2 antibodies in the serum of patients with hepatitis C virus (HCV) infection.

Methods

The microplate was coated with recombinant envelope proteins E1 and E2 by indirect method, respectively, and the kits for detecting anti-E1 and anti-E2 antibodies were prepared. The methodological indexes were evaluated.

Results

The methodological indexes of the kits were as follows: precision test (the variation coefficient of anti-E1 antibody 6.71%–8.95% for within run and 9.91%–12.16% for between run, the variation coefficient of anti-E2 antibody 6.06%–8.44% for within run and 10.77%–13.98% for between run, respectively). The blank limit and detection limit were 1.18 RLIR and 3.16 RLIR for the anti-E1 antibody, and 1.26 RLIR and 3.32 RLIR for the anti-E2 antibody, respectively. The correlation coefficients (r) of anti-E1 and anti-E2 were 0.9963 and 0.9828, the analysis and measurement ranges (AMR) were 1.66–41.28 RLIR and 1.55–19.46 RLIR, and the average recovery was 96.4% and 93.7%, respectively. The rheumatoid factor and other positive serum samples had no interference or cross-reaction to the test, and the kits were stable within 15 months. The positive rates of anti-E1 and anti-E2 antibodies in 45 patients with HCV infection were 35.6% (16/45) and 44.4% (20/45), respectively.

Conclusions

The kits for detecting anti-E1 and anti-E2 meet the requirements of methodology, and can be used in screening diagnosis, disease monitoring, prognosis evaluation, disease mechanism, and epidemiological studies of HCV infection. The HCV envelope proteins E1 and E2 have an immune response in HCV-infected patients.

背景建立一种检测丙型肝炎病毒（HCV）感染者血清中抗 E1 和抗 E2 抗体的化学发光方法。方法采用间接法在微孔板上分别涂布重组包膜蛋白 E1 和 E2，制备检测抗 E1 和抗 E2 抗体的试剂盒。对方法学指标进行了评价。结果试剂盒的方法学指标如下：精密度试验（抗-E1 抗体的运行内变异系数为 6.71%-8.95%，运行间变异系数为 9.91%-12.16%；抗-E2 抗体的运行内变异系数为 6.06%-8.44%，运行间变异系数为 10.77%-13.98%）。抗 E1 抗体的空白限和检测限分别为 1.18 RLIR 和 3.16 RLIR，抗 E2 抗体的空白限和检测限分别为 1.26 RLIR 和 3.32 RLIR。抗 E1 和抗 E2 的相关系数（r）分别为 0.9963 和 0.9828，分析和测量范围（AMR）分别为 1.66-41.28 RLIR 和 1.55-19.46 RLIR，平均回收率分别为 96.4% 和 93.7%。类风湿因子和其他阳性血清样本对检测无干扰或交叉反应，试剂盒在 15 个月内性能稳定。45 名 HCV 感染者的抗 E1 和抗 E2 抗体阳性率分别为 35.6%（16/45）和 44.4%（20/45）。结论抗 E1 和抗 E2 检测试剂盒符合方法学要求，可用于 HCV 感染的筛查诊断、疾病监测、预后评估、疾病机制和流行病学研究。HCV 包膜蛋白 E1 和 E2 在 HCV 感染者中具有免疫反应。

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引用次数: 0

Exploration of the Clinical Effect of Different Autotransfusion Methods on Patients With Femoral Shaft Fracture Surgery 探讨不同自体输血方法对股骨柄骨折手术患者的临床效果。

IF 2.7 4区医学 Q2 MEDICAL LABORATORY TECHNOLOGY

Journal of Clinical Laboratory Analysis

Pub Date : 2024-03-11 DOI: 10.1002/jcla.25018

Yujia Wang, Huan Wang, Jiaming Xu, Jinhuo Wang, Laiwei You, Yu Bai, Jianrong Guo

Objective

To explore the clinical effect of predeposit, salvage, and hemodilution autotransfusion on patients with femoral shaft fracture (FSF) surgery.

Methods

Selected patients with FSF were randomly divided into three groups: intraoperative blood salvage autotransfusion, preoperative hemodilution autohemotransfusion, and predeposit autotransfusion. Five days after the operation, the body temperature, heart rate, blood platelet (PLT), and hemoglobin (Hb) of patients were determined. The concentrations of EPO and GM-CSF in the three groups were calculated by ELISA. The content of CD14+ monocytes was calculated by FCM assay. The growth time and condition of the patient's callus were determined at the 30th, 45th, and 60th day after operation. Cox regression analysis was used to analyze the correlation between EPO, GM-CSF, CD14+ mononuclear content, callus growth, and autotransfusion methods.

Results

There were no statistically significant differences in body temperature and heart rate between the three groups (p > 0.05). PLT and Hb in the Predeposit group were markedly increased compared with that in the Salvage and Hemodilution groups. The concentrations of EPO and GM-CSF in the Predeposit group were markedly increased compared with that in the Salvage and Hemodilution groups. The content of CD14+ monocytes in the Predeposit group was significantly higher than that in the Salvage and Hemodilution groups. Predeposit autotransfusion promotes callus growth more quickly.

Conclusion

Predeposit autotransfusion promoted the recovery of patients with FSF after the operation more quickly than salvage autotransfusion and hemodilution autotransfusion.

目的探讨股骨干骨折（FSF）手术患者术前、术中、术后三组自体输血的临床效果：将选定的股骨干骨折患者随机分为三组：术中血液抢救性自体输血组、术前血液稀释性自体输血组和预存款自体输血组。术后五天，测定患者的体温、心率、血小板（PLT）和血红蛋白（Hb）。用酶联免疫吸附法计算三组患者体内 EPO 和 GM-CSF 的浓度。通过 FCM 检测法计算 CD14+ 单核细胞的含量。在术后第 30 天、第 45 天和第 60 天测定患者胼胝体的生长时间和状况。采用 Cox 回归分析法分析 EPO、GM-CSF、CD14+ 单核细胞含量、胼胝体生长和自体输血方法之间的相关性：三组体温和心率差异无统计学意义（P > 0.05）。与抢救组和血液稀释组相比，预存组的 PLT 和 Hb 明显增加。与抢救组和血液稀释组相比，预置组的 EPO 和 GM-CSF 浓度明显升高。预处理组 CD14+ 单核细胞的含量明显高于抢救组和血液稀释组。结论：结论：与抢救性自体输血和血液稀释性自体输血相比，预置自体输血能更快地促进FSF患者术后恢复。

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引用次数: 0

IGF2BP2 and IGFBP3 Genotypes, Haplotypes, and Genetic Models Studies in Polycystic Ovary Syndrome 多囊卵巢综合征的 IGF2BP2 和 IGFBP3 基因型、单倍型和遗传模型研究。

IF 2.7 4区医学 Q2 MEDICAL LABORATORY TECHNOLOGY

Journal of Clinical Laboratory Analysis

Pub Date : 2024-03-11 DOI: 10.1002/jcla.25021

Fatemeh Govahi Kakhki, Saman Sargazi, Farzaneh Montazerifar, Mahdi Majidpour, Atena Karajibani, Mansour Karajibani, Marzieh Ghasemi

Background

Insulin resistance has been correlated with the genetic diversity within the insulin-like binding proteins genes. Moreover, insulin resistance is one of the key characteristics of the widespread reproductive endocrine condition known as polycystic ovarian syndrome (PCOS). Hence, this study is aimed to determine the association between IGFBP3 and IGF2BP2 gene variants and PCOS risk.

Methods

A total of 300 subjects (150 PCOS cases diagnosed based on Rotterdam ESHRE/ASRM consensus criteria and 150 healthy subjects) were recruited in this case–control cross-sectional study. Tetra-primer amplification refractory mutation system polymerase chain reaction (ARMS-PCR) was used for genotyping rs11705701, whereas genotyping of rs1470579 and rs2854744 was done employing PCR-restriction fragment length polymorphism (PCR-RFLP) technique.

Results

The CC and AA+AC genotypes of rs1470579 conferred an increased risk of PCOS in our population. Regarding the rs2854744, an increased risk of PCOS was observed under the codominant homozygous (TT vs. GG) model by 2.54 fold. The C allele of rs1470579 and T allele of rs2854744 enhanced PCOS risk by 1.97 and 1.46 folds, respectively. Haplotype analysis showed that the A_rs1470579A_rs11705701 haplotype conferred a decreased risk of PCOS (odds ratio = 0.53, 95% confidence interval = 0.34–0.83, p = 0.006). The AC/GG/GT, AA/GA/GT, AC/GA/GG, and AC/GA/GT genotype combinations of rs1470579/rs11705701/rs2854744 were associated with a decreased risk of the disease.

Conclusions

IGF2BP2 rs1470579 and IGFBP3 rs2854744 enhanced PCOS susceptibility in a Southeastern Iranian population. Further investigation involving larger cohorts representing diverse ethnic backgrounds is needed to confirm the current findings.

背景：胰岛素抵抗与胰岛素样结合蛋白基因的遗传多样性有关。此外，胰岛素抵抗是被称为多囊卵巢综合征（PCOS）的广泛生殖内分泌疾病的主要特征之一。因此，本研究旨在确定 IGFBP3 和 IGF2BP2 基因变异与多囊卵巢综合征风险之间的关联：这项病例对照横断面研究共招募了 300 名受试者（150 名根据鹿特丹 ESHRE/ASRM 共识标准确诊的 PCOS 病例和 150 名健康受试者）。rs11705701的基因分型采用四引物扩增难治性突变系统聚合酶链反应（ARMS-PCR），rs1470579和rs2854744的基因分型采用PCR-限制性片段长度多态性（PCR-RFLP）技术：结果：在我们的人群中，rs1470579的CC和AA+AC基因型会增加患多囊卵巢综合征的风险。至于 rs2854744，在共显同型（TT 与 GG）模式下，多囊卵巢综合征的风险增加了 2.54 倍。rs1470579 的 C 等位基因和 rs2854744 的 T 等位基因分别将 PCOS 风险提高了 1.97 倍和 1.46 倍。单倍型分析表明，Ars1470579 Ars11705701 单倍型可降低 PCOS 风险（几率比 = 0.53，95% 置信区间 = 0.34-0.83，p = 0.006）。rs1470579/rs11705701/rs2854744的AC/GG/GT、AA/GA/GT、AC/GA/GG和AC/GA/GT基因型组合与患病风险降低有关：结论：在伊朗东南部人群中，IGF2BP2 rs1470579 和 IGFBP3 rs2854744 会增加多囊卵巢综合症的易感性。要证实目前的研究结果，还需要对更多不同种族背景的人群进行进一步调查。

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引用次数: 0