Nondestructive fluorescence-based quantification of threose-induced collagen cross-linking in bovine articular cartilage

ABSTRACT. Extensive collagen cross-linking affects the mechanical competence of articular cartilage: it can make the cartilage stiffer and more brittle. The concentrations of the best known cross-links, pyridinoline and pentosidine, can be accurately determined by destructive high-performance liquid chromatography (HPLC). We explore a nondestructive evaluation of cross-linking by using the intrinsic fluorescence of the intact cartilage. Articular cartilage samples from bovine knee joints were incubated in threose solution for 40 and 100?h to increase the collagen cross-linking. Control samples without threose were also prepared. Excitation-emission matrices at wavelengths of 220 to 950?nm were acquired from the samples, and the pentosidine and pyridinoline cross-links and the collagen concentrations were determined using HPLC. After the threose treatment, pentosidine and lysyl pyridinole (LP) concentrations increased. The intrinsic fluorescence, excited below 350?nm, decreased and was related to pentosidine [r=-0.90, 240/325 nm (excitation/emission)] or LP (r=-0.85, 235/285 nm) concentrations. Due to overlapping, the changes in emission could not be linked specifically to the recorded cross-links. However, the fluorescence signal enabled a nondestructive optical estimate of changes in the pentosidine and LP cross-linking of intact articular cartilage.     

Level of agreement on postoperative complications after one-stage hypospadias correction comparing medical records and parent reports

PurposeTo analyze agreement on postoperative complications after hypospadias surgery according to medical records and parents' reports.Materials & methodsIn this retrospective cohort study, data were collected from 409 children who received an initial one-stage hypospadias correction in the Radboudumc, The Netherlands.Postoperative complications according to medical records were compared with parent-reported complications in an online questionnaire. Main complications studied were wound-related complications, urinary tract infections, fistulas, stenosis, and prepuce-related complications. Agreement was determined by Cohen's kappa coefficient.ResultsSlightly less complications were mentioned in medical records (37%) compared to parents' reports (42%). Overall agreement was moderate (κ = 0.50, 95% confidence interval (CI):0.41–0.59), but poor for some specific complications. Agreement was higher for complications that needed reoperation compared to when no reoperation was performed (κ = 0.53, 95% CI: 0.43–0.62 and κ = 0.18, 95% CI: 0.06–0.31) and for patients with recent surgery (< 5 years before questionnaire completion) compared to less recent surgeries (κ = 0.69, 95% CI: 0.55–0.84 and κ = 0.43, 95% CI: 0.33–0.54).ConclusionsAgreement on complications according to medical records and parents' reports was poor to moderate, but better after reoperation and more recent surgery. Some complications mentioned in medical records were missing from parents' reports and the other way around. Better agreement will give physicians and parents a more reliable view on postoperative outcome after hypospadias surgery.Type of studyDiagnostic test.Level of evidenceLevel III.     

KGF-2与PRO2605蛋白相互作用的初步研究

目的：利用酵母双杂交系统筛选KGF-2的相互作用蛋白，为阐明其分子作用机制提供有益线索。方法：通过聚合酶链反应从人胎肝cDNA文库中钓取KGF-2 cDNA，构建诱饵蛋白载体pAS2-1-KGF-2，并对其自身转录激活活性进行鉴定，利用酵母双杂交系统筛选人胎肝cDNA文库，挑选双阳性克隆。将KGF-2和候选蛋白分别克隆至哺乳动物细胞双杂交的BD、AD质粒中，共同转染COS-7细胞，通过CAT分析验证KGF-2和候选蛋白之间的相互作用。结果：VDNA序列分析和同源检索显示所获候选蛋白为PR02605。以KGF-2和PRO2605 cDNA共转化酵母宿主Y190后可激活报道基因，但共转染COS-7细胞后，CAT分析结果阴性。结论：KGF-2和PRO2605在酵母体内存在相互作用，但在COS-7细胞中未能检测到两者的相互作用。