A comparative study of P53/MDM2 genes alterations and P53/MDM2 proteins immunoreactivity in soft-tissue sarcomas

In the present study, the expression of P53 and MDM2 proteins were examined in 94 soft-tissue sarcomas (35 malignant fibrohistiocytomas, 15 neurosarcomas, 14 liposarcomas, 13 leiomyosarcomas, 11 fibrosarcomas and 6 dermatofibrosarcomas) by immunohistochemistry. The immunohistochemical findings were correlated with P53 mutation analysis using PCR-SSCP, PCR-HDF and direct sequencing, and MDM2 amplification studies by differential PCR. P53 immunopositivity was found in 25 out of 94 (26.6%) cases. Alterations of the P53 gene were detected in 12 (12.8%) tumors; eight of these tumors revealed P53 immunoreactivity. A high number of P53 positive and P53 mutated tumors were histologically defined as poorly differentiated G3 (64.0% and 75.0%, respectively). MDM2 immunopositivity was revealed in 36 out of 94 (38.3%) cases. MDM2 amplification occurred in 17 tumors (18.1%); only nine of these tumors exhibited MDM2 immunoreactivity. Overall, MDM2 positivity was not associated with MDM2 amplification in 27 out of 94 tumors (28.7%). There was no significant correlation between MDM2 overexpression and histological grade. However, when the samples were stratified by immunophenotype, the majority of tumors (52.5%) with isolated MDM2 overexpression (dissociated from P53 positivity) were defined histologically as low grade (G1 + G2). These results support the notion that besides P53 alterations, MDM2 gene deregulation seems to be an important event in sarcomas evolution. Additionally, the mechanism of MDM2-mediated degradation of P53 protein, without involving stabilization and inactivation of P53 gene, should be considered for better understanding of all features of tumor progression processes.     

Polyelectrolyte microcapsules with entrapped multicellular tumor spheroids as a novel tool to study the effects of photodynamic therapy

Our purpose of this study is to develop potassium or sodium/strontium co-substituted calcium polyphosphate (K/Sr-CPP or Na/Sr-CPP) bioceramics in application of bone repairing scaffold. The incorporation of K, Na, and Sr into CPP substrate via a calcining-sintering process was confirmed by X-ray diffractometry and inductively coupled plasma atomic emission spectroscopy. In vitro degradation study of co-substituted CPP indicated the incorporation of alkali metal elements promoted the degradability of CPP, and the scanning electron microscope showed the apatite-like minerals were precipitated on the surface of co-substituted CPP. The compress resistant strength of co-substituted CPP was elevated by dopants. The MTT assay and confocal laser-scanning microscope on osteoblasts culturing with co-substituted CPP showed no cytotoxicity. The cell proliferation on co-substituted CPP was even better than others. Thus, this co-substituted CPP bioceramics might have potential of applications in orthopedic field.     

A New Locus for Familial FSGS on Chromosome 2P

FSGS is a clinicopathologic entity characterized by nephrotic syndrome and progression to ESRD. Although the pathogenesis is unknown, the podocyte seems to play a central role in this disorder. Here, we present six kindreds with hereditary FSGS that did not associate with mutations in known causal genes, and we report a new locus for the disease on chromosome 2p15 in one kindred. We performed genome-wide linkage analysis and refined the linkage area with microsatellite markers and haplotype analysis to define the minimal candidate region. Genome-wide linkage analysis yielded a maximum two-point logarithm of odds (LOD) score of 3.6 for the six families on chromosome 2p. One family contributed the largest proportion of the additive score (LOD 2.02) at this locus. Multipoint parametric LOD score calculation in this family yielded a significant LOD score of 3.1 at markers D2S393 and D2S337, and fine mapping of this region with microsatellite markers defined a minimal candidate region of 0.9 Mb with observed recombinations at markers D2S2332 and RS1919481. We excluded the remaining five families from linkage to this region by haplotype analysis. These data support a new gene locus for familial FSGS on chromosome 2p15. Identification of the mutated gene at this locus may provide further insight into the disease mechanisms of FSGS.FSGS is a clinicopathologic entity that is characterized by the nephrotic syndrome and is often steroid resistant. Progression to end-stage kidney disease (ESKD) is frequent. Histologically, the lesion is characterized by focal glomerulosclerosis or tuft collapse, segmental hyalinosis, occasionally IgM staining on immunofluorescence, and effacement of foot processes on electron microscopy.¹ Its incidence is estimated at seven per million.² It is responsible for 2 to 20% of all cases of ESKD in the United States, and it is second only to urogenital and kidney malformation as a cause of ESKD in children.^2–5 In addition, it is the most common glomerular cause of ESKD.³ The incidence of FSGS seems to be increasing. Kitiyakara et al.⁵ reported an 11-fold increase among dialysis patients who were older than 21 years. In every age group, the incidence is higher in black than in white patients with a striking difference in the age distribution pattern in black patients; the highest incidence occurred in the 40- to 49-year age group.^2,5,6FSGS can be broadly classified into primary (idiopathic), secondary, syndromic, and familial. Familial cases are increasingly being recognized. The primary defect in FSGS seems to be in the filtration barrier of the glomerulus. This barrier is made up of the fenestrated endothelium, the glomerular basement membrane, and podocytes that have a slit diaphragm between their interdigitating foot processes. Disruption of the filtration barrier results in loss of permselectivity and albuminuria. The pathogenesis of FSGS has yet to be fully explained. Early theories included T lymphocyte dysfunction and dysregulation of cytokines and growth factors.^7,8 Recent results from genetic studies reveal mutations in genes that encode slit diaphragm and podocyte cytoskeletal proteins. There is now strong evidence to suggest that FSGS is essentially a disease of the podocyte. The first major breakthrough was the cloning of nephrin (NPHS1) as a cause of congenital nephrotic syndrome of the Finnish type.⁹ Subsequently, five additional genes have been identified, including podocin (NPHS2); actinin-α4 (ACTN4/FSGS1); transient receptor potential cation channel, type 6 (TRPC6/FSGS2); CD2-associated protein (CD2AP/FSGS3); and phospholipase c, ε1 (PLCE1/NPHS3) as causes for FSGS and hereditary nephrotic syndromes.^10–14The discovery of these genes has allowed us to begin to unravel signaling events and cytostructural abnormalities in the podocyte and the slit diaphragm. Further studies of familial cases will help in understanding the fundamental pathways for the ultrastructural and functional changes seen in FSGS and the subsequent development of rational approaches to therapy. In addition, because the pathologic findings in the more common sporadic FSGS are similar to those of inherited FSGS, it is possible that common mechanisms are responsible for disease evolution in both familial and sporadic cases. Despite the advance in knowledge regarding inherited FSGS, there remains a subset of families who have hereditary FSGS but do not carry known disease-causing mutations.We present a comprehensive genetic study of six kindreds with familial FSGS, which includes the sequencing of known FSGS genes and genes for other hereditary nephrotic syndromes, genome-wide linkage analysis, fine-mapping, and haplotype analysis. There is no evidence for linkage or mutations in previously identified genes in these families. Genome-wide linkage analysis identified three regions of linkage. Concentrated analysis of one very large family supported strong linkage to one of these regions, and fine-mapping narrowed the linkage region to a 0.9-Mb interval on chromosome 2p15.     

A Review of the Website TeamScience.net

As modern research methods have become more specialized and the true complexity of today’s most pressing health issues and diseases is revealed, collaborations among scientists trained in different fields have become essential for exploring and tackling these problems. This specialization of research methods has made interdependence, joint ownership, and collective responsibility between and among scientists near requirements. These features of team science may not suit everyone, but given these current trends, it is increasingly likely that most researchers will find themselves asked to participate on or lead a research team at some point in their careers.Bennett, Gadlin, Levine-Finley¹     

The interplay of HER2/HER3/PI3K and EGFR/HER2/PLC-γ1 signalling in breast cancer cell migration and dissemination

HER2 signalling by heterodimerisation with EGFR and HER3 in breast cancer is associated with worst outcome of the afflicted patients, which is attributed not only to the aggressiveness of such tumours but also to therapy resistance. Thus, in the present study we investigated the role of EGFR, HER2 and HER3 lateral signalling in cell migration by applying the MDA-MB-468-HER2 (MDA-HER2) breast cancer cell line, representing a valid model system. Knockdown of HER3 expression by siRNA resulted in decreased phosphorylated AKT (pAKT) levels, abrogated epidermal growth factor (EGF)-mediated PLC-γ1 activation and a diminished EGF-induced migratory activity, depicting the interplay of EGF receptor (EGFR)/HER2/PLC-γ1 and HER2/HER3/PI3K signalling in mediating the migration of EGFR/HER2/HER3-expressing breast cancer cells. Since therapy failure usually arises from metastatic cells, we further investigated whether HER3 signalling was active in established breast cancer disseminated tumour cell (DTC) lines as well as in primary DTCs derived from breast cancer patients. EGF treatment of DTC lines resulted solely in increased pAKT S473 levels, whereas in MDA-HER2 cells both pAKT S473 and pAKT T308 levels were increased upon EGF stimulation. Moreover, despite active HER3 molecules, as indicated by pTyr1222 staining, about 90% of analysed breast cancer patient DTCs exhibited very low or even no detectable pAKT S473 levels, suggesting that these cells might have fallen into dormancy. In summary, our data indicate the important role in EGFR, HER2 and HER3 lateral signalling in breast cancer cell migration. Moreover, our data further show that primary tumour cells and DTCs can vary in their HER activation status, which is important to know in the context of cancer therapy.     

Circulating tumour cell-derived plastin3 is a novel marker for predicting long-term prognosis in patients with breast cancer

Background:

Identification of promising biomarkers that predict the prognosis of patients with breast cancer is needed. In this study, we hypothesised that the expression of the epithelial–mesenchymal transition-related biomarker plastin3 (PLS3) in peripheral blood could be a prognostic factor in breast cancer.

Methods:

We examined PLS3 expression in breast cancer cell lines with epithelial and mesenchymal traits and in circulating tumour cells (CTCs) obtained from the peripheral blood of breast cancer patients. We investigated PLS3 expression in the peripheral blood of 594 patients with breast cancer to evaluate the clinical significance of PLS3 expression.

Results:

Robust PLS3 expression was observed in different breast cancer cell lines (Hs578t, MCF-7, MDA-MB-468, and MDA-MB-231) as well as in a bone marrow derived cancer cell line (BC-M1). In both the training (n=298) and validation (n=296) sets, PLS3 expression was observed in CTCs of patients with breast cancer. PLS3-positive patients showed significantly poorer overall and disease-free survival than PLS3-negative patients (P=0.0001 and 0.003, respectively). Subset analysis revealed that this prognostic biomarker was relevant in patients with stage I–III cancer, particularly in patients with luminal-type and triple-negative-type tumours.

Conclusions:

These data demonstrated that PLS3 was expressed in CTCs undergoing the epithelial–mesenchymal transition in patients with breast cancer. Furthermore, PLS3 may be an excellent biomarker for identifying groups at risk of recurrence or with a poor prognosis.     

Inverted formin 2 mutations with variable expression in patients with sporadic and hereditary focal and segmental glomerulosclerosis

Focal and segmental glomerulosclerosis (FSGS) is a major cause of end-stage kidney disease. Recent advances in molecular genetics show that defects in the podocyte play a major role in its pathogenesis and mutations in inverted formin 2 (INF2) cause autosomal dominant FSGS. In order to delineate the role of INF2 mutations in familial and sporadic FSGS, we sought to identify variants in a large cohort of patients with FSGS. A secondary objective was to define an approach for genetic screening in families with autosomal dominant disease. A total of 248 individuals were identified with FSGS, of whom 31 had idiopathic disease. The remaining patients clustered into 64 families encompassing 15 from autosomal recessive and 49 from autosomal dominant kindreds. There were missense mutations in 8 of the 49 families with autosomal dominant disease. Three of the detected variants were novel and all mutations were confined to exon 4 of INF2, a regulatory region responsible for 90% of all changes reported in FSGS due to INF2 mutations. Thus, in our series, INF2 mutations were responsible for 16% of all cases of autosomal dominant FSGS, with these mutations clustered in exon 4. Hence, screening for these mutations may represent a rapid, non-invasive and cost-effective method for the diagnosis of autosomal dominant FSGS.