Diagnostic usefulness of EMA, IMP3, and GLUT-1 for the immunocytochemical distinction of malignant cells from reactive mesothelial cells in effusion cytology using cytospin preparations

To differentiate reactive mesothelial cells (RMs) from metastatic carcinoma and malignant mesothelioma (MM) in effusion cytology is crucial for the cytologic diagnosis and the management of the patients. In the present study, the immunocytochemical staining profile of the epithelial membrane antigen (EMA), the insulin-like growth factor-II mRNA-binding protein 3 (IMP3), and the glucose transporter-1 (GLUT-1) was examined to distinguish RMs from malignant cells. A total of 171 pleural (n = 87) and peritoneal (n = 84) effusion specimens, including 50 benign effusions with RMs, 11 MM effusions, and 110 metastatic malignant effusions, were evaluated for immunocytochemistry. EMA, IMP3, monoclonal GLUT-1, and polyclonal GLUT-1 immunoreactivity were observed in 26.0%, 6.0%, 20.0%, and 18.0% of RMs, respectively. In contrast to RMs, the immunoreactivity in MM was 100%, 36.4%, 100%, and 90.9%; adenocarcinoma (AC) was 100%, 80.8%, 81.7%, and 72.1%; squamous-cell carcinoma was 83.3%, 83.3%, 83.3%, and 66.7%. EMA, IMP3, mGLUT-1, and pGLUT-1 expressions were observed in 98.4%, 65.6%, 88.5%, and 75.4% in the pleural effusion with malignant cells, and 100%, 88.3%, 78.3%, and 71.7% in ascites containing malignant cells, respectively. The findings of the present study indicate that the immunocytochemical staining for EMA, IMP3, and GLUT-1 is a useful diagnostic tool for distinguishing effusions containing malignant cells from those that contain benign cells, and in particular, we suggest that the combination of mGLUT-1 and EMA, and IMP3 and EMA are extremely useful in pleural effusion and in ascites, respectively.     

Utility of ProExC and IMP3 immunocytochemical staining in atypical glandular cells of undetermined significance in liquid‐based cervical cytology

The diagnosis of atypical glandular cells of undetermined significance (AGUS) in liquid‐based cervical cytology specimens shows significant underlying pathology in only 30% of cases, while the remaining cases are found to be benign (reactive, reparative/metaplastic). Previous studies have reported positive ProExC and IMP3 staining in neoplastic glandular lesions of the uterine cervix and corpus. We present our experience with the utility of these markers in the evaluation of AGUS cases in liquid‐based cervical cytology. The case cohort included 34 cases diagnosed as AGUS. ProExC and IMP3 immunocytochemical (ICC) stains were performed on ThinPrep® slides and the results correlated with subsequent biopsy findings. Positive expression was classified as strong diffuse nuclear immunostaining for ProExC and granular cytoplasmic for IMP3. The presence of AGUS cells on the ICC stained slides was confirmed in all cases. IMP3 was positive in 80% of glandular neoplasms and negative in 93% non‐glandular lesions/cases negative for squamous intraepithelial lesion (SIL). ProExC was positive in 60% of glandular neoplasms and negative in 83% non‐glandular lesions/cases negative for SIL. When used as a panel (ProExC + IMP3), at least one stain was positive in 100% of glandular neoplasm cases and they were both negative in 83% of non‐glandular lesions/cases negative for SIL. Based on this study, both ProExC and IMP3, when used as an immuno panel, can predict the presence of glandular lesions on subsequent biopsies and can serve as an aid in the diagnosis and management of AGUS cases. Diagn. Cytopathol. 2014;42:375–379. © 2013 Wiley Periodicals, Inc.     

Role of lymphovascular invasion and immunohistochemical expression of IMP3 in the risk stratification of superficially invasive pT1 esophageal adenocarcinoma

Background

A problem in the management of patients with Barrett's esophagus-related pT1 esophageal adenocarcinoma is to distinguish those who should be treated conservatively (endoscopic mucosal resection and/or radiofrequency ablation) from those who require esophago-gastrectomy. Recently, lymphovascular invasion (LVI) has emerged as one of the best predictors of regional lymph node metastasis (LNM) and recurrence-free survival (RFS) in pT1 EAC. However, LVI may be underestimated, both because of interobserver variability and incomplete sampling. The aim of our study was to correlate the presence of LVI, with the immunohistochemical expression of IMP3 in pT1 EAC and assess their role in further stratifying these lesions into high and low risk groups based on the potential for lymph node metastasis and poor outcome.

Design

Depth of invasion, assessed in five sublevels (m2, m3, sm1, sm2, and sm3), LVI, and expression of IMP3 were studied in 30 patients who underwent esophagogastrectomy for pT1 EAC (2001–2010) at Hartford Hospital, and correlated with LNM and RFS. IMP3 was considered positive when expressed in >50% of the malignant cells with an intensity of stain of 2–3+.

Results

Ten of 18 (55.5%) cases with IMP3 expression demonstrated LVI and 2/10 (20%) showed LNM and died of disease. In contrast, none of the 12 IMP3 negative cases showed LVI (p < 0.004; 2-tailed Fisher exact test) or had LNM/DOD.

Conclusions

In pT1 EAC, (1) based on IMP3 expression, pT1 EAC may be divided into high risk (LVI+/IMP3+) and low risk (LVI−/IMP3−) categories. (2) Absence of IMP3 expression is associated with a significantly reduced risk of LVI (Negative Predictive Value: 100%). (3) Since identifying lymphovascular invasion and other morphological parameters is prone to significant inter-observer variation, IMP3 may be useful as an ancillary marker especially in these pT1 lesions in predicting their clinical behavior, the risk stratification and potentially on the type of treatment.     

The expression of IMP3 in 366 cases with ovarian carcinoma of high grade serous,endometrioid and clear cell subtypes

The clear cell (CCC), high grade serous (HGSC) and endometrioid (EC) ovarian carcinomas share overlapping histological features. The oncogene IMP3 is implicated in CCC with an elusive utility in differential diagnosis. We collected 366 cases with ovarian primary carcinomas to detect IMP3, Napsin-A and HNF-1β by immunochemistry. In 351 cases, the positive expression rate of IMP3 in CCC was significantly higher than that either in EC or HGSC (p?<?0.01). The sensitivity of IMP3 in CCC was higher than Napsin-A but lower than HNF-1β (p?<?0.01). The specificity of IMP3 in CCC was lower than Napsin-A but higher than HNF-1β (p?<?0.01). The composite markers Napsin-A+/IMP3+ and the IMP3+/HNF-1β+/Napsin-A+ offered the highest odds ratio (p?<?0.001), the highest specificity, the highest positive predictive value and the highest positive likelihood ratio. The ROC analysis showed that the combination of Napsin-A, HNF-1β and IMP3 offered the biggest AUC compared with either the singular marker performances or the other binary combinations (p?<?0.001). In 15 cases of EC mixed with CCC, IMP3 showed a better discrimination value than the other two markers. Consequently, adding IMP3 to the diagnostic panel might provide some help with the pathological diagnosis of ovarian CCC.     

A fine decision tree consisted of CK5/6, IMP3 and TTF1 for cytological diagnosis among reactive mesothelial cells,metastatic adenocarcinoma of lung and non-lung origin in pleural effusion

The utility of combination with CK5/6, IMP3 and TTF1 to differentiate among reactive mesothelial cells (RMs), metastatic adenocarcinoma of lung (LAC) and non-lung (NLAC) origin was investigated by using immunocytochemistry (ICC) and conventional PCR (C-PCR) in pleural effusion. A total of 108 cell blocks (32 RMs, 51 LAC and 25 NLAC were evaluated by ICC for CK5/6, IMP3 and TTF1 protein expression. In addition, we further performed C-PCR for amplification of CK5/6, IMP3 and TTF1 DNA from 28 specimens (9 MAC and 7 RMs, 6 LAC and 6 NLAC) for molecular diagnosis. CK5/6 staining was observed in the majority of reactive specimens (78.1%) and was rare in adenocarcinoma cells (14.5%), whereas the opposite was true for IMP3 and TTF1. We found a high frequency of TTF1 positivity (76.5%) in LAC, but not in NLAC (4.0%); while there was no significant difference of IMP3 expression in LAC (88.2%) and NLAC (88.0%). The 487 bp DNA fragments of IMP3 was expected to be amplified in 6/9 of adenocarcinoma cases showed negative in ICC; and the 394 bp DNA fragments of CK5/6 was also expected to be amplified in 4/7 of RMs cases showed negative in ICC. Consistent with ICC results, there was significant difference of TTF1 expression in the LAC and NLAC compared with IMP3 expression. The combination with CK5/6, IMP3 and TTF1 immunostaining appears to be useful to improve the accuracy of cytological diagnoses between RMs, metastatic adenocarcinoma of lung and non-lung origin in pleural effusion. In addition, C-PCR may act as a useful supplemental approach for ICC, especially negative cases in ICC for differential cytological diagnosis.     

Utility of pVHL,maspin, IMP3, S100P and Ki67 in the distinction of autoimmune pancreatitis from pancreatic ductal adenocarcinoma

Morphology plays an important role in the distinction of autoimmune pancreatitis (AIP) from pancreatic ductal adenocarcinoma (PDAC). However, we aimed to determine the utility of immunohistochemical tumor markers to contribute in the distinction of these entities. In surgical specimens with AIP (n = 20), PDAC (n = 20) and normal pancreas (n = 20), the expression of pVHL, maspin, IMP3, S100P and Ki67 was examined. We evaluated intralobular reactive ducts / acinoductal metaplasia (ILDs) and extralobular ducts (ELDs) in AIP, neoplastic glands in PDAC, and ductal epithelium in the normal pancreas, using a five-tiered scoring system. The Ki67 hot spot index (Ki67-HSPI) was determined manually and using automated digital imaging analysis of virtual double stains of Ki67 and CK8. Besides, sequential dual-immunohistochemical staining of maspin/pVHL, maspin/IMP3 and Ki67/maspin was performed in a subset of the specimens. Strong overexpression of IMP3, maspin, S100P and Ki67 and loss of pVHL was observed in PDAC compared to AIP and normal pancreas. In AIP however, focal and weak aberrant expression was observed with the following proportions in ILDs/ELDs: pVHL in 45 %/85 %, maspin in 30 %/70 %, IMP3 in 55 %/5%, S100P in 10 %/35 % and Ki67-HSPI >20 % in 15 %/70 %. At least two markers were aberrantly expressed in ILDs/ELDs in 45 %/60 %. The aberrant expression was more pronounced in type 2 AIP compared to type 1. In conclusion, our data indicate that pVHL, maspin, IMP3, S100P and Ki67 can be focal and weak aberrantly expressed in AIP. However, when used as a panel, these markers seem to be useful for the differentiation of AIP from PC.     

Distinguishing benign from malignant mesothelial cells in effusions by Glut‐1, EMA,and Desmin expression: An evidence‐based approach

Distinguishing malignant mesothelioma (MM) from reactive mesothelial hyperplasia (RM) may be difficult in effusions. This study tested the hypothesis that immunocytochemistry (IC) in effusion cell blocks (CB) can distinguish MM from RM and that the results may be applied to individual specimens. External validation of a risk score (RS) model associating sensitivity and specificity was applied to an external set of MM and RM specimens from a separate institution. Forty three effusion cytology CBs of 25 confirmed malignant mesotheliomas were compared to CBs of 23 benign mesothelial effusions without inflammation and 13 reactive mesothelial proliferations associated with inflammation. Glut‐1, EMA, and Desmin expression were evaluated by immunocytochemistry on CBs. Each antibody was compared using ROC values, where the area under the curve (AUC) was 0.90, 0.82, and 0.84 for Glut‐1, EMA, and Desmin, respectively. Logistic regression (LR) analysis was applied to a combination of Glut‐1 and EMA. A combined ROC curve was modeled for Glut‐1 and EMA (AUC = 0.93). A RS = 2 × (Glut‐1%) + 1 × (EMA%) was created from this ROC curve. When applied to an external set of MM and RM, the RS resulted in an ROC with AUC = 0.91. In conclusion, a RS derived from a LR of Glut‐1 and EMA IC greatly improves the distinction between MM from RM cells in individual effusions. The study illustrates principles of evidence‐based pathology concerning internal and external test performance in the differential diagnosis of MM versus RM. Diagn. Cytopathol. 2013. © 2011 Wiley Periodicals, Inc     

Distribution and significance of 14-3-3sigma, a novel myoepithelial marker, in normal, benign, and malignant breast tissue

14-3-3sigma is a candidate tumour suppressor gene transactivated by p53 in response to DNA damage. In gene expression analysis of normal luminal and myoepithelial cells, 14-3-3sigma was preferentially expressed by myoepithelial cells. This study has analysed the immunohistochemical distribution and subcellular localization of 14-3-3sigma in normal breast tissue and in a large series of benign and malignant breast lesions on whole tissue sections and by tissue microarray. Immunohistochemistry demonstrated that 14-3-3sigma was consistently expressed in the cytoplasmic compartment and occasionally in the nuclei of myoepithelial cells arranged as a continuous layer around normal ducts and lobular units, whereas luminal epithelial, stromal, endothelial, pericytic, lipomatous, and neural cells showed no staining. Myoepithelial cells of benign proliferations and pre-invasive lesions were consistently positive for 14-3-3sigma. Strong expression of 14-3-3sigma was evident in one case of ductal carcinoma in situ (5.5%) and in 105/554 invasive cancers (18.9%). Survival data were available for 452 patients with invasive breast carcinoma. 14-3-3sigma cytoplasmic subcellular localization was a statistically significant prognostic factor for the whole series of invasive carcinomas, as well as for those positive for oestrogen (ER) or progesterone receptors (PR). This analysis demonstrates the utility of 14-3-3sigma as a new adjunct antibody for characterization of myoepithelial cells and myoepithelial lesions and it may be a novel prognostic factor for breast cancer patients.     

Epithelial membrane antigen in the cytodiagnosis of effusions and aspirates: immunocytochemical and ultrastructural localization in benign and malignant cells

Immunoreactivity for epithelial membrane antigen (EMA) was evaluated in exfoliated benign and malignant cells using immunoperoxidase and immunogold techniques. In addition, protein A-colloidal gold was used for ultrastructural localization of EMA. Our results suggest that EMA is useful in distinguishing adenocarcinoma cells (strongly positive) from reactive mesothelial cells (negative or weakly positive), lymphoid cells (negative), and a variety of nonepithelial neoplasms (negative) with which they may be confused. Exfoliated cells from two mesotheliomas were also strongly positive for EMA. Ultrastructurally, EMA was distributed in a dense, even, linear pattern along the cell membrane and microvillous surface processes of adenocarcinoma cells. A similar but sparse distribution pattern was observed in reactive mesothelial cells. These observations reflect the increased sensitivity and higher resolution of the immunogold technique.     

Utility of thin-layer preparations in endometrial cytology: immunocytochemical expression of PTEN, beta-catenin and p53 for benign endometrial lesions

This article focuses on the characteristic features of morphology and molecular biology of PTEN, beta-catenin, and p53 immunocytochemistry in normal endometrium (proliferative, secretory, and atrophic) and endometrial glandular and stromal breakdown (EGBD) using thin-layer specimens. During a 6-month period, 120 endometrial samples were collected directly using the Uterobrush and a thin-layer specimen was prepared. Immunocytochemical expression of PTEN, beta-catenin, and p53 were investigated using 30 cases each of proliferative endometrium (PE), secretory endometrium (SE), atrophic endometrium (AE), and EGBD.PTEN expression of normal endometrial glandular epithelial cells changes with the hormonal status; PE produce very high expression, SE creates attenuation or disappearance of PTEN expression and AE diminished more in comparison with SE. PTEN expression of EGBD showed a tendency towards attenuation compared with PE, but showed high expression in comparison with SE and AE. As for the immunoreactivity of beta-catenin, in all phases (PE, SE, AE, and EGBD), it was observed in the cytoplasm of glandular epithelial cells, but not nuclei, and showed strong membranous staining. As for the p53 immunoreactivity, p53 positivity was not observed in the glandular epithelial cells in all phases (PE, SE, AE, and EGBD) with the exception of some metaplastic cells. The presence of p53 immunoreactivity of a weak, low ratio in metaplastic cells was unexpected. In the current study, the expression manner of PTEN, beta-catenin, and p53 immunocytochemistry was observed in the normal endometrium (PE, SE, and AE) and EGBD.     

High expression of cytoplasmic phosphorylated CSE1L in malignant melanoma but not in benign nevi: phosphorylated CSE1L for the discrimination between melanoma and benign nevi

Melanoma is difficult to treat when it has metastasized. Discrimination between melanoma and benign nevi in melanocytic lesions is crucial for identifying melanomas and consequently improving melanoma treatment and prognosis. The chromosome segregation 1-like (CSE1L) protein has been implicated in cancer progression and is regulated by phosphorylation by extracellular signal-regulated kinase 1/2 (ERK1/2) signaling, a critical pathway in melanoma progression. We studied phosphorylated CSE1L expression in human melanoma and benign nevi specimens. Immunohistochemistry with tissue microarray using antibody against phosphorylated CSE1L showed that melanomas exhibited considerable staining for phosphorylated CSE1L (100%, 34/34), whereas the benign nevi showed only faint staining (0%, 0/34). Melanomas mainly exhibited cytoplasmic phospho-CSE1L distribution, whereas the benign nevi mainly exhibited nuclear phospho-CSE1L distribution. Moreover, immunohistochemistry with anti-CSE1L antibody revealed that CSE1L mainly exhibited cytoplasmic/nuclear distribution and nuclear distribution was the dominant. Immunofluorescence with B16F10 melanoma cells showed cytoplasmic distribution of phospho-CSE1L and nuclear distribution of CSE1L. Our results indicated that nuclear CSE1L is mainly non-phosphorylated CSE1L and is involved in gene regulation and cytoplasmic CSE1L is mainly phosphorylated CSE1L and is involved in cytoplasmic signaling regulation in melanocytic tumorigenesis. Furthermore, immunohistochemical analysis of cytoplasmic phospho-CSE1L may aid in the diagnosis of melanoma.     

Galectin-3、CK19和TPO表达在甲状腺良恶性肿瘤病理诊断中的价值

目的比较甲状腺病变中galectin-3、CK19和甲状腺过氧化物酶（thyroid peroxidase,TPO）蛋白表达的差异,寻找有助于诊断良恶性肿瘤的标志物。方法采用免疫组化EnVision二步法检测134例甲状腺癌（123例乳头状癌、11例滤泡癌）、34例甲状腺腺瘤、20例结节性甲状腺肿和10例桥本甲状腺炎中galectin-3、CK19和TPO蛋白的表达。结果Galectin-3、CK19和TPO在甲状腺乳头状癌（包括主要的3个亚型）与良性病变中的表达差异有显著性（P〈0．01）,在乳头状癌各亚型间和各良性病变之间的表达差异均无显著性（P〉0．05）。滤泡癌和良性病变中galectin-3表达差异有显著性（P〈0．01）。滤泡癌和腺瘤中CK19、TPO的表达差异有显著性（P〈0．05,P〈0．01）。结论甲状腺恶性肿瘤特别是乳头状癌中galectin-3和CK19的表达有增高,而TPO表达缺失。3种蛋白分别是诊断甲状腺肿瘤特别是乳头状癌的有用标志物,联合使用结果更可靠。     

Activation of the PI3K/Akt/mTOR/p70S6K Pathway is Involved in S100A4-induced Viability and Migration in Colorectal Cancer Cells

The S100 protein family member S100A4 regulates various cellular functions. Previous studies have shown that elevated expression of S100A4 is associated with progression and metastasis of colorectal cancer (CRC). However, little is known about whether and how S100A4 contributes to CRC development. In our present study, the elevated expression of S100A4 in CRC tissues compared to matched adjacent normal tissues was confirmed by immunohistochemistry, semi-quantitative RT-PCR and Western blot. Adenovirus-mediated S100A4 overexpression obviously enhanced viability and migration of CRC cells, which was detected by MTT assay and transwell assay, respectively. Additionally, S100A4 overexpression increased the phosphorylation levels of Akt, mTOR and p70S6K. These effects of S100A4 were abolished by treatment with either the specific PI3K/Akt inhibitor {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}}LY294002, or the specific mTOR/p70S6K inhibitor rapamycin. Furthermore, overexpression of S100A4 resulted in upregulation of VEGF and downregulation of E-cadherin, which were strongly reversed by either {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}}LY294002 or rapamycin. Altogether, our results demonstrate that activation of the PI3K/Akt/mTOR/p70S6K signaling pathway is involved in S100A4-induced viability, migration, upregulation of VEGF and downregulation of E-cadherin in CRC cells.     

The process of granule exocytosis in non-stimulated atrial granular cells of the snail, Achatina achatina: An ultrastructural,histochemical and immunocytochemical study

Abundant secretory granular cells (GCs) in the Giant African land snail atrium harbor a range of bioactive substances and undergo rapid total degranulation in response to stimulation of the cardiac nerve or stressful influences. Here we have analyzed exocytotic events in the non-stimulated GCs. It was shown that the GCs contain three major distinct types of granules that differ histochemically, immunocytochemically and ultrastructurally, each performing specific functions. The type I granules characteristically filled with electron-lucent homogeneous materials exhibit intense immunoreactivity for bioactive proteins and therefore are considered to be storage granules. Histochemistry using vital staining with Acridine Orange and Gomori acid phosphatase technique has revealed lysosomal-related nature of the electron-dense type II granules. Digestion remnants appearing as fine filamentous materials fill the type III granules. Only the type III granules fuse together and with the plasma membrane form degranulation channels and surface pores, through which the debris is removed from the cell. The finding of granules exhibiting intermediate ultrastructural, histochemical and immunocytochemical features suggests that the major granule types represent most stable states along a granule empting continuum. Thus, under physiological conditions, the GCs continuously produce secretory proteins and so maintain readiness for stress-response, but use protein degradation machinery to prevent massive release of these bioactive substances into hemolymph.     

Identification of mammosomatotropes, growth hormone cells and prolactin cells in the pituitary gland of the gilthead sea bream (Sparus aurata L., Teleostei) using light immunocytochemical methods: an ontogenetic study

Growth hormone (GH) and prolactin (PRL) immunoreactivities in the adenohypophysis of Sparus aurata specimens from newly hatched until 48-months-old were detected using the peroxidase-antiperoxidase method. GH cells and PRL cells, and cells that were immunoreactive to both GH and PRL antisera, called mammosomatotropes (MS cells), were found. This is the first report on the identification of MS cells in fish, which were found in newly hatched and older larvae and juvenile specimens. GH and PRL cells appeared from two days after hatching. MS cells were first located in the central region of the adenohypophysis and afterwards in the rostral pars distalis. The GH cells were first identified in the dorsal and ventral areas of the middle-posterior part, and the PRL cells in the ventral region of the middle-anterior part. Later, during development, the sequence of appearance of the GH cells was proximal pars distalis, pars intermedia and rostral pars distalis, while for the PRL cells sequence was rostral pars distalis, proximal pars distalis and pars intermedia. This expansion pattern could be due to a GH- and PRL-cell migration although independent cell differentiation may occur in each region. The present results suggest that GH and PRL cells arise from MS cells at the outset of pituitary development, while MS cells procede from PRL cells in old larvae and juveniles. Accepted: 8 June 2000     

Metabolic activation of 3-methylcholanthrene and benzo(a)pyrene to mutagens in the L5178Y/TK assay by cultured embryonic rodent cells

Early passage cultured rodent cells obtained from mouse, rat, or Syrian hamster embryos were cocultivated with L5178Y/TK^+/- cells in the presence of benzo(a)pyrene (B(a)P) or 3-methylcholanthrene (3MCA) for 6 hr to determine if active metabolic intermediates could be detected in this cell-mediated assay through the induction of TK^+/- mutants. Neither chemical was directly active as a mutagen when tested with L5178Y/TK^+/- cells alone. As a mutagen, 3MCA was slightly active in the presence of mouse or rat embryo cells and clearly active when tested with hamster embryo cells. B(a)P was active only in the presence of hamster embryo cells when tested at concentrations that would be mutagenic in the presence of rodent liver microsomal preparations. These results indicate the utility of L5178Y/TK^+/- plus hamster embryo cell combinations in a cell-mediated mutation assay for the detection of these two polycyclic hydrocarbon carcinogens.     

Overexpression of FER1L4 promotes the apoptosis and suppresses epithelial-mesenchymal transition and stemness markers via activating PI3K/AKT signaling pathway in osteosarcoma cells

Novel long non-coding RNA Fer-1-like protein 4 (FER1L4) has been identified as a tumor suppressor in endometrial carcinoma, ovarian cancer, hepatocellular carcinoma, esophageal squamous cell carcinoma. However, the function of FER1L4 in osteosarcoma has not been clear. The aim of the research was to explore the effects of FER1L4 in osteosarcoma. Results showed that FER1L4 was observed to be lowly expressed in osteosarcoma cell lines (US-O2, MG-63 and SaOS-2 cells), especially MG63 cells. Besides, overexpression of FER1L4 remarkably repressed the proliferation, migration and invasion of MG63 cells. FER1L4-induced apoptotic cell death leaded to the activation of caspase-3 and Bax/Bcl2. Moreover, epithelial-mesenchymal transition (EMT) was tremendously suppressed by increased FER1L4, evidences were the increased E-cadherin and reduced vimentin and fibronectin. Blocking FER1L4 expression by sh-FER1L4 treatment increased the expression of SOX9, CD44, ALDH1, Nanog and Oct4, indicating that FER1L4 could effectively decrease cell stemness in osteosarcoma. Furthermore, the protein levels of p-AKT and p-PI3K were remarkably suppressed when FER1L4 was knocked down. In conclusion, the study indicated that FER1L4 acted as a tumor suppressor in osteosarcoma via activating PI3K/AKT pathway may be a new prognostic biomarker and potential therapeutic target for osteosarcoma intervention.     

Early onset and slow progression of SCA28, a rare dominant ataxia in a large four-generation family with a novel AFG3L2 mutation

Autosomal dominantly inherited spinocerebellar ataxias (SCAs) are a heterogeneous group of neurodegenerative disorders primarily affecting the cerebellum. Genetically, 26 different loci have been identified so far, although the corresponding gene has not yet been determined for 10 of them. Recently, mutations in the ATPase family gene 3-like 2 gene were presented to cause SCA type 28. To define the frequency of SCA28 mutations, we performed molecular genetic analyses in 140 unrelated familial cases with ataxia. Among other variations, we found a novel missense mutation at an evolutionarily conserved amino-acid position using a single-strand conformation polymorphism approach, followed by DNA sequencing. This amino-acid exchange p.E700K was detected in a four-generation German family and was not observed in a survey of 400 chromosomes from healthy control individuals.