Desmin-free cardiomyocytes and myocardial dysfunction in end stage heart failure

Our aim was to evaluate the desmin content in the myocardial tissue of patients with end-stage heart failure of ischaemic origin and to assess its role on cardiac function. We studied 18 explanted hearts from patients transplanted for end-stage heart failure due to ischaemic cardiomyopathy (ICM). Control myocardial tissue was obtained from the cardiac biopsies of six women with breast cancer taken prior to commencing chemotherapy with anthracyclines, four male donors for heart transplantation and two autoptic hearts from patients who died due to non-cardiac events. Myocardial tissue, obtained from the left ventricle (remote zone from infarcted area), was analyzed by light and confocal immunochemistry (desmin) microscopy. The desmin content of myocardial tissue was obtained by real-time PCR. Cardiac function was evaluated by echocardiographic and right heart catheterization data, obtained before heart transplantation. Confocal microscopy evaluation showed a significant decrease in the number of desmin-positive myocytes (P<0.01) in ICM hearts compared to controls. At real-time PCR evaluation, there was a reduction (P<0.01) in desmin content in the ICM patients compared to controls. A negative correlation was found between desmin-free cardiomyocytes and ejection fraction (EF) (r=-0.834; P<0.02) on echocardiogram. A negative relationship (r=-0.688) was also found between desmin-negative myocytes and capillary wedge pressure. In conclusion, the myocardial tissue of patients with end-stage heart failure of ischaemic origin, shows a decreased number in desmin-positive myocytes at immunochemistry evaluation compared to normal individuals. This deficiency in cytoskeletal intermediate filament content is associated with reduced cardiac function.     

N-乙酰半胱氨酸对糖尿病大鼠肾脏的保护作用

目的 观察抗氧化剂N－乙酰半胱氨酸（NAC）对糖尿病大鼠肾脏保护作用. 方法 14只大鼠单次腹腔注射链脲佐菌素（STZ） 60 mg&#8226;kg^-1制备糖尿病模型,随机分为糖尿病组和治疗组各7只,另设正常组7只. 治疗组给予NAC 125 mg&#8226;kg^-1&#8226;d^-1,8周后测3组大鼠的肾重/体重,血糖（BS）、24 h尿清蛋白定量 （UAE）、三酰甘油（TG）、胆固醇（TC）、内生肌酐清除率（Ccr）;并测肾皮质内丙二醛（MDA）含量和总抗氧化能力（TAOC）. PAS染色和电镜观察肾小球结构,免疫组化检测肾小球内结蛋白（desmin）表达水平. 结果 与糖尿病组比较,治疗组肾重/体重、UAE,TC,Ccr都显著下降;肾皮质内MDA含量显著降低,而TAOC显著升高. 治疗组肾小球病理结构和电镜结构较糖尿病组有所改善. Desmin免疫组化显示正常组肾小球内有少量表达,糖尿病组肾小球内表达显著高于治疗组. 结论 NAC可以延缓糖尿病肾损害进展,其作用机制可能是通过减轻肾组织氧化应激和足细胞损伤所致.     

冻椎间盘复合表达hBMP7的髓核细胞体外构建生物椎间盘的可行性研究

目的 探讨利用液氮储存的椎间盘与表达人BMP7的髓核细胞体外构建生物活性椎间盘器官的可行性.方法 取液氮储存2个月的犬椎间盘48个,分为EGFP对照组、1×10^4、1×10^5和1×106共四组,每组12个椎间盘.对照组用22 G注射器自椎间盘后正中注入20 μL 含有1×10^5表达EGFP的髓核细胞,1×10^4组注入20 μL含有1×10^4 PKH-26标记的表达hBMP7的髓核细胞,1×10^5组注入20 μL含1×106KH-26标记的表达hBMP7的髓核细胞,1×106组注入20 μL含1×10^5KH-26标记的表达hBMP7的髓核细胞.注射后立即置入50 mL离心管中,加入30 mL完全培养基,分别于培养第4、7、14 d,从外源细胞存活、存活细胞量、hBMP7的表达、蛋白多糖及总胶原含量变化对体外构建的生物活性椎间盘进行评价.结果 在椎间盘培养的各时间点均可见PKH-26红色荧光和表达绿色荧光蛋白的髓核细胞存在.对不同组、不同时间点的椎间盘中荧光强度定量后发现：培养第7天、14天时1×10^5组荧光强度明显高于1×106组和1×10^4组（P〈0.05）.1×10^5组在第7、14天时hBMP7 mRNA表达量明显高于1×106组和1×10^4组（P〈0.05）,EGFP组未见表达;且1×10^5组在培养的第7天、14天时蛋白多糖及总胶原含量均较另外三组明显增高（P〈0.05）.结论 应用液氮冻存的椎间盘与1×10^5个表达hBMP7的髓核细胞复合,外源性髓核细胞可较长时间存活,并能增加蛋白多糖及胶原含量,是一种体外构建生物活性椎间盘的可行性方法.     

Right ventricular remodeling in restrictive ventricular septal defect

Restrictive ventricular septal defect (rVSD) presents with little/no hemodynamic aberrations despite a patent septal defect. Clinically, these patients are observed with the hope that the defect will functionally close over time without the need for surgical repair and development of heart failure. Without evidence supporting a definitive therapeutic strategy, rVSD patients may have increased risk of a poor outcome. We tested the hypothesis that rVSD results in subclinical RV diastolic dysfunction and molecular remodeling. Five pigs underwent surgical rVSD creation. Echocardiography, hemodynamics, myocyte contractility experiments, and proteomics/Western blot were performed 6-weeks post-rVSD and in controls. *p < 0.05. LV and RV hemodynamics in rVSD were comparable to controls. The tricuspid valve early/late diastolic inflow velocity ratio (TV E/A ratio) decreased from 1.6 ± 0.05 in controls to 1.0 ± 0.08* in rVSD, indicating RV diastolic dysfunction. rVSD RV myocytes showed abnormalities in contraction (departure velocity (Vd) − 51%*, Vd time + 55%*) and relaxation (return velocity (Vr) − 50%*, Vr time + 62%*). Mitochondrial proteins (fatty acid, TCA cycle) increased 2-fold*, indicating heightened RV work. Desmin protein upregulated 285%* in rVSD RV myocardium, suggesting cytoskeletal remodeling. rVSD causes RV diastolic dysfunction, myocyte functional impairment, and mitochondrial/cytoskeletal protein upregulation in our model. Desmin upregulation may hinder sarcomeric organization/relaxation, representing a key subclinical early marker for future RV dysfunction. TV E/A measurements are a non-invasive modality to assess rVSD patients for diastolic dysfunction. Translational research applications may lead to fundamental changes in the clinical management of rVSD by providing evidence for early repair of the defect.     

The effect of vitamin A on CCl4-induced hepatic injuries in rats: a histochemical, immunohistochemical and ultrastructural study

In this study, we aimed to investigate the effect of vitamin A on the transformation of the Ito cells to fibrogenic form and suppression of the development of fibrosis. Carbon tetrachloride intoxication was performed on rats for 2, 8, 12 or 20 weeks and 5x10(4) IU vitamin A (as retinol palmitate) was injected subcutaneously once every 4 weeks. Ito cells were detected by gold chloride impregnation, as well as desmin and alpha-smooth muscle actin (alpha-SMA) immunohistochemistry. Additionally, all groups were examined ultrastructurally. The number of Ito cells that were labelled positively with gold impregnation decreased in the fibrotic groups; however, alpha-SMA and desmin immunopositive Ito cells increased. The samples from animals that were treated with vitamin A showed an increase in labelling with gold impregnation but a decrease in alpha-SMA immunopositivity. The data showed that vitamin A can prevent hepatic injury, by suppressing the transformation of Ito cells to fibrogenic form. We conclude that vitamin A has potential for the treatment of hepatic fibrotic diseases. Alpha-SMA immunohistochemistry was found to be more informative than desmin immunohistochemistry for monitoring liver fibrosis.     

南宁地区动脉内膜结蛋白与动脉粥样硬化的观察

连续１２例意外死亡的广西自治区南宁市年轻人（１５～３４岁）免疫组化染色证明，其主动脉及冠状动脉内膜的平滑肌细胞含有较多结蛋白。南宁是我国动脉粥样硬化最低发区之一，也是世界低发区。结蛋白阳性是平滑肌细胞成熟的标志。成熟平滑肌细胞为收缩型细胞，不易受生长因子作用而活跃增殖。因而认为这可能是动脉粥样硬化低发区的特征。与北京相比较，本地区动脉壁含较多具有抗增殖活性的硫酸乙酰肝素蛋白聚糖。平滑肌细胞产生这种聚糖构成细胞外层，像筑茧自缚一样抑制自身生长，因而平滑肌细胞比较成熟稳定，显示较多结蛋白。     

Time-dependent appearance of myofibroblasts in granulation tissue of human skin wounds

Summary Human skin wounds (66) inflicted between 20 h and 7 months prior to biopsy were studied. In order to identify the type of cellular differentiation of the fibroblastic cells in the granulation tissue, alpha-smooth muscle actin and desmin were immunohistochemically localized. The value of any presumed time-dependent appearance and/or disappearance of positively stained cells was tested for the estimation of wound age. In skin specimens with a wound age less than 5 days (n =15) no typical granulation tissue had developed and no alpha-actin-positive myofibroblasts could be detected. The first appearance of positively reacting myofibroblasts was noted in a 5-day-old wound. In 57% of the lesions with a wound age between 5 and 31 days (25 out of 44 cases) typical granulation tissue formation was present and myofibroblasts with positive reaction for alpha-smooth muscle actin could be identified. Numerous positively reacting cells could generally be found in wounds aged between 16 and 31 days, but also in wounds less than 16 days old. In 29% of the cases with a wound age of more than 31 days (2 out of 7 cases) alpha-sma-positive myofibroblasts also occured. Fibroblastic cells positive for desmin could not be seen at all in our series. Our results demonstrate the appearance of alpha-sma-positive myofibroblasts with the initial formation of typical granulation tissue in human skin lesions as early as approximately 5 days after wounding. In contrast to recent experimental results these cells remained detectable in wounds aged more than 2 months in some cases. The immunohistochemical detection of actin-positive cells, therefore, demonstrates whether an unknown skin wound is aged approximately 5 days or more. Even though a time-dependent decrease of myofibroblasts in human granulation tissue after 31 days in human wounds seems probable, the extended presence (up to about 2 months) of these cells allows no further exact age determination of older wounds. This study was supported by a grant from the “Deutsche Forschungsgemeinschaft” (grant Ei 209/3-1) and by a grant from the “Friedrich-Baur-Stiftung”, University of Munich, FRG.     

Experimental emetine myopathy: enzyme histochemical,electron microscopic,and immunomorphological studies

Summary Ipecac, containing emetine hydrochloride, is used by patients with anorexia nervosa to induce vomiting. Its chronic usage may result in a myopathy and a cardiomyopathy, the former marked by cytoplasmic bodies. We studied myopathological changes after daily injections of female Wistar rats with emetine hydrochloride intraperitoneally for periods of 4, 5, 9, and 10 weeks. the extensor digitorum longus muscle and the soleus muscle showed core-like lesions, streaming of the z-discs, nemaline bodies, cytoplasmic bodies, and spheroid cytoplasmic bodies. Immunomorphological studies revealed increased amounts of desmin. During a period of repair, i.e., 2, 4, and 6 weeks after termination of emetine application, these myopathological alterations faded while proliferation of the T-tubular system, i.e., honeycomb structures, was more often prevalent. Pathological features completely disappeared between 6 and 12 weeks of recovery.     

Neuromuscular damage after hyperthermic isolated limb perfusion in patients with melanoma or sarcoma treated with chemotherapeutic agents

Purpose: To evaluate the incidence and entity of muscle damage after hyperthermic limb perfusion (HLP) with doxorubicin or melphalan, two widely used chemotherapeutic agents. Methods: We collected muscle biopsies from eleven patients with lower limb sarcoma or melanoma immediately before and at a variable time after the chemotherapeutic procedure (mean= 49.4 days). Biopsy specimens were stained with standard histochemical and immunohistochemical methods on cryostat sections and the grade of fiber atrophy was calculated. Results: Clear neurogenic alterations were present in pre-HLP biopsies of seven patients related to age and previous therapy. In six patients, the comparison between biopsies before and after HLP demonstrated worsening of preexisting neurogenic condition and appearance of mitochondrial-related damage. Reduction in type I or type II fiber diameter was present in nine patients, but no relation to doxorubicin or melphalan treatment was clear. An unexpected, large accumulation of desmin was detected in the muscle biopsy of one patient receiving doxorubicin, probably related to the mechanism of doxorubicin-induced myotoxicity. Conclusions: The observed neuromuscular toxic effects could be related to the physical or chemical conditions of HLP, in particular perfusion temperature; in addition, the present study demonstrates that preexisting neuromuscular changes, i.e. neuropathy, modulates the degree of further damage following HLP. Received: 16 March 2000 / Accepted: 26 June 2000     

子宫内膜间质肉瘤特异性标志物的研究

目的 :探讨子宫内膜间质肉瘤的特异性标志物。方法 :选取子宫内膜间质肉瘤 2 6例 (低度恶性 2 1例 ,高度恶性 5例 ) ,普通型和富于细胞平滑肌瘤各 2 5例 ,正常子宫肌层和正常子宫内膜各 2 5例 ,用免疫组化SP法检测各种组织中雌、孕激素受体 ,结蛋白 ,调宁蛋白 (calponinh1 ) ,以及CD1 0和CD44v3的表达。结果 :子宫内膜间质肉瘤和富于细胞平滑肌瘤均表达雌、孕激素受体和结蛋白。调宁蛋白在所有正常子宫肌层和平滑肌瘤中强表达 ,在 9例子宫内膜间质肉瘤中亦有表达。所有正常子宫内膜间质和 2 1例低度恶性子宫内膜间质肉瘤均弥漫强表达CD1 0 ,而富于细胞平滑肌瘤中仅 1例局灶表达。富于细胞平滑肌瘤中均可见CD44v3表达 ,而在子宫内膜间质肉瘤中仅 2例表达 ,用CD44v3缺如诊断子宫内膜间质肉瘤的敏感性为 92 .3 %,特异性达 1 0 0 %。结论 :CD1 0是低度恶性子宫内膜间质肉瘤较特异的指标。调宁蛋白和CD44v3可用来辅助鉴别子宫内膜间质肉瘤和富于细胞平滑肌瘤