Actin structural proteins in cell motility

Summary The machinery for cell locomotion is based in a network of polymerized actin filaments supporting the peripheral cytoplasm. This network or gel consists of actin filaments in a variety of configurations, including cables, loose bundles, and branching arrays; all formed by the interaction of actin-associated proteins with actin filaments. For cell locomotion to occur, this network must be reversibly disassembled or solated to allow protrusion, then re-assembled to stabilize the resulting extension. Thus, proteins to promote both solation and gelation of actin are important for efficient cell locomotion. Because of their distribution, control, and in vitro effects on actin filaments, two such proteins, gelsolin and actin-binding protein (ABP) should play especially important roles in cell motility. Support for this premise is found in in vivo studies of mouse kidney fibroblasts which demonstrated increased translocational locomotion after cytoplasmic gelsolin expression was increased genetically and in melanoma cells missing actin-binding protein which behave as expected for a cell unable to achieve efficient actin gelation. Since malignant transformation is known to affect the expression and distribution of several of these actin structural proteins, including gelsolin, further investigations of the role these proteins play in cell motility will be important to the determination of tumor cell motility and hence metastatic propensity.     

Synergistic antineoplastic action of DNA methylation inhibitor 5-AZA-2'-deoxycytidine and histone deacetylase inhibitor depsipeptide on human breast carcinoma cells

During tumorigenesis, cancer-related genes can be silenced by aberrant DNA methylation and by changes in chromatin structure. It has been reported that 5-aza-2'-deoxycytidine, a potent inhibitor of DNA methylation, in combination with histone deacetylase inhibitors, can produce a synergistic reactivation of these genes. The aim of our study was to investigate the in vitro antineoplastic activity of 5-aza-2'-deoxycytidine in combination with depsipeptide, a potent histone deacetylase inhibitor, against MDA-MB-231 and MDA-MB-435 human breast carcinoma cell lines. We observed that the combination of 5-aza-2'-deoxycytidine and depsipeptide produced a synergistic antineoplastic effect against these tumor cells as compared to either agent administered alone. We also investigated the effect of this drug combination on the activation of maspin and gelsolin expression. These 2 genes whose function is to suppress tumor metastasis have been reported to be silenced by epigenetic events in breast cancer. Using semi-quantitative RT-PCR, we observed that 5-aza-2'-deoxycytidine in combination with depsipeptide produced a greater reactivation of both maspin and gelsolin as compared to each agent alone. The synergistic interaction between 5-aza-2'-deoxycytidine and depsipeptide on breast carcinoma cell lines provides a rationale to investigate this interesting drug combination in future clinical trials on patients with advanced breast cancer.     

Downregulation of Gelsolin Correlates with the Progression to Breast Carcinoma

The actin cytoskeleton underlies several normal cellular functions and is deranged during carcinogenesis. Gelsolin, a multifunctional actin-binding protein, is downregulated in several types of tumors and its abnormal expression is one of the most common defects noted in invasive breast carcinoma (ICA). This study utilizes immunohistochemistry to examine the expression of gelsolin in 95 ICA, 59 ductal carcinoma in situ (DCIS) and 36 benign lesions, including 17 atypical ductal hyperplasia (ADH). Cytoplasmic staining was scored as positive, reduced or negative. Gelsolin expression was then correlated with patients age, tumor size, histologic grade and lymph node status. All unremarkable breast biopsies, 88% of ADH, 44% of DCIS and 28% of ICA were positive for gelsolin. This represents a significant difference among the groups (p=<0.0001) and the trend towards reduced gelsolin with the progression to ICA is significantly linear (p=<0.0001). For invasive carcinoma, patients older than 44 years were significantly more likely to have decreased expression of gelsolin than patients 44 years old and younger (p=0.007). Bivariate analysis showed no correlation of gelsolin expression with lymph node status (p=0.62), tumor size (p=0.10), histologic grade (p=0.42), estrogen receptor status (p=1.0) or other clinicopathologic parameters. In clinical follow-up, there were 18 breast tumor related deaths within a median follow-up time of 4.2 years. Survival analysis indicated that the level of gelsolin expression may be associated with survival (p=0.06). In summary, the frequency of gelsolin deficiency increases significantly with progression from ADH to DCIS to ICA. Additionally, gelsolin expression may be an independent marker of prognosis.     

Lattice corneal dystrophy,gelsolin type (Meretoja’s syndrome)

Purpose: This paper reviews current knowledge about the pathogenesis, clinical manifestations and treatment of lattice corneal dystrophy, gelsolin type (LCD2, Meretoja’s syndrome). Methods: Material is derived from literature searches, a case study of a Finnish patient living in Sweden, and interviews in Helsinki with Professor Ahti Tarkkanen and Dr Sari Kiuru‐Enari, both of whom have extensive first‐hand experience in treating patients with the disease. Results: The disease is now reported from several countries in Europe, as well as Japan, the USA and Iran. Treatment is symptomatic and is based on eye lubrication combined with rigorous monitoring of intraocular pressure to reduce corneal haze and postpone the need for keratoplasty. When systemic symptoms occur, the ophthalmologist should consult other specialists. Conclusions: The disease is probably under‐reported and is almost certainly to be found in more countries, including Sweden. Every ophthalmologist should be vigilant and consider this diagnosis when discovering a corneal lattice dystrophy, especially because the disease is an inherited, lifelong chronic condition with systemic symptoms.     

Proteomics of tumor-specific proteins in cerebrospinal fluid of patients with astrocytoma: Usefulness of gelsolin protein

Changes in cerebrospinal fluid (CSF) composition have been shown to accurately reflect pathological processes in the CNS, and are potential indicators of abnormal CNS states, such as tumor growth. To detect biomarkers in high-grade astrocytomas, the differential expression of proteins in the cerebrospinal fluid was analyzed from two cases each of diffuse astrocytoma (grade II), and glioblastoma (grade IV) using agarose 2-D gel electrophoresis (2-DE). It was found that the expression of gelsolin protein decreased with histological grade. To examine whether gelsolin is a useful indicator of tumor aggressiveness or patient outcome, its expression was further studied on immunohistochemistry in 41 formalin-fixed and paraffin-embedded astrocytomas. The positive cell rate of gelsolin in tumors was 59.4% in grade II, 30.0% in grade III and 29.4% in grade IV, respectively. Gelsolin expression was significantly lower in high-grade astrocytomas (grade III or IV) than in low-grade astrocytomas (grade II; P < 0.05). Moreover, in astrocytomas the overall survival of patients in the low-expression group was significantly poorer than in the high expression group ( P < 0.05). These data suggest that gelsolin is a prognostic factor in astrocytoma.     

Frequencies and geographic distributions of genetic mutations in transthyretin‐ and non‐transthyretin‐related familial amyloidosis

Inherited forms of amyloidosis are rare; of these, transthyretin‐related (ATTR) is the most common, but non‐ATTR has been described as well. We studied a large case series of ATTR and a small series of non‐ATTR to better determine the mutation frequencies and geographic distributions of these inherited forms of amyloidosis in the United States. We performed a retrospective cross‐sectional study of 284 ATTR and non‐ATTR patients seen at Mayo Clinic in Rochester, Minnesota, from 1 January 1970 through 29 January 2013. Mutations were identified by DNA sequencing, restriction fragment length polymorphism, or mass spectroscopy. The genetic testing method was unknown for several patients, but a small proportion were identified by family history or by classical clinical presentation associated with a specific mutation. The most common ATTR mutations were Thr60Ala (24%), Val30Met (15%), Val122Ile (10%), and Ser77Tyr (5%). Non‐ATTR mutations included gelsolin (n = 3), apolipoprotein A‐I (n = 6), apolipoprotein A‐II (n = 1), fibrinogen A‐α (n = 9), and lysozyme (n = 1). Although rare, ATTR and, to a lesser extent, non‐ATTR are prevalent in the United States and should be considered for patients presenting in the appropriate clinical context.     

凝溶胶蛋白抑制乳腺癌细胞MCF7／Her2增殖及迁移侵袭的作用

目的研究凝溶胶蛋白（gelsolin,GSN）对雌激素受体α（ERα）阳性、人表皮生长因子受体2（Her2）过表达乳腺癌细胞MCF7／Her2生长的作用。方法构建GSN的真核表达载体,并稳定转染MCF7／Her2细胞,以RT-PCR及WesternBlot鉴定得到稳定过表达GSN的乳腺癌细胞系McF7／Her2／GsN。以空载体转染的稳定表达细胞MCF7／Her2／V作对照,比较其细胞形态、增殖和迁移能力的改变;WesternBlot及RT-PCR检测Her2、组织金属蛋白酶抑制剂-3（TIMP3）表达和细胞外调节蛋白激酶（ERK）的活化,探索深层次的作用机制。结果MCF7／Her2稳定过表达GSN后,细胞形态改变且增殖及迁移、侵袭能力明显低于对照细胞;发现Her2表达下降,TIMP3的表达上调,而雌激素导致的MCF7／Her2细胞ERK活化增强被逆转。结论凝溶胶蛋白能降低Her2的表达,并通过下调ERK的活化及上调TIMP3的表达,抑制乳腺癌细胞MCF7／Her2增殖及迁移侵袭作用。     

Bacterial endotoxin as inhibitor of the enzymatic activity of human thrombin

Endotoxemia caused by bacterial lipopolysaccharides (LPS) deleteriously affects many aspects of hemostasis. Much of this effect is well characterized as being secondary to the LPS-mediated inflammatory response, but direct effects of LPS on coagulation factors may also contribute to disregulation of the hemostatic process. Spectrophotometric assays were used to investigate the effects of LPS from different bacteria on thrombin and plasmin activities. We found that enzymatic activity of purified thrombin, but not plasmin, decreases in the presence of endotoxin. LPS-mediated inhibition of thrombin activity can be reversed by plasma gelsolin and recombinant endotoxin-neutralizing protein. Preincubation of thrombin with LPS before platelet activation results in inhibition of aggregation and secretion. Additionally, a decrease of elastic shear moduli of fibrin gels was observed when their formation was induced with thrombin preincubated with LPS or when LPS was present in fibrinogen solutions during fibrin gel formation. When added to platelet-rich plasma, after activation with collagen, LPS-inhibited thrombin activity. LPS-mediated inhibition of thrombin activity may contribute to the hemostasis dysfunctions observed during endotoxemia.