The Interplay Between Autophagy and RNA Homeostasis: Implications for Amyotrophic Lateral Sclerosis and Frontotemporal Dementia.

Amyotrophic lateral sclerosis and frontotemporal dementia are neurodegenerative disorders that lie on a disease spectrum, sharing genetic causes and pathology, and both without effective therapeutics. Two pathways that have been shown to play major roles in disease pathogenesis are autophagy and RNA homeostasis. Intriguingly, there is an increasing body of evidence suggesting a critical interplay between these pathways. Autophagy is a multi-stage process for bulk and selective clearance of malfunctional cellular components, with many layers of regulation. Although the majority of autophagy research focuses on protein degradation, it can also mediate RNA catabolism. ALS/FTD-associated proteins are involved in many stages of autophagy and autophagy-mediated RNA degradation, particularly converging on the clearance of persistent pathological stress granules. In this review, we will summarise the progress in understanding the autophagy-RNA homeostasis interplay and how that knowledge contributes to our understanding of the pathobiology of ALS/FTD.

A rare case of retroperitoneal synovial sarcoma.

Management of retroperitoneal soft tissue sarcomas is complex. Treatment is usually multimodal; involving surgery, chemotherapy and radiotherapy.

Morules in endometrioid proliferations of the uterus and ovary consistently express the intestinal transcription factor CDX2.

AIMS: To undertake an immunohistochemical analysis of squamous elements in endometrioid proliferations of the uterus and ovary and to compare the immunophenotype of typical squamous elements and so-called squamous morules. METHODS AND RESULTS: Cases of uterine or ovarian endometrioid glandular lesions with squamous elements were stained with CDX2, beta-catenin, oestrogen receptor (ER), CD10, p63 and high-molecular-weight cytokeratin LP34. Thirteen cases had typical squamous elements and 18 cases morules. Morules typically exhibited diffuse nuclear CDX2 and beta-catenin immunoreactivity and were positive for CD10 and LP34. They were usually ER- and p63-. In contrast, typical squamous elements were usually positive for ER, CD10, p63 and LP34. They were usually CDX2- or focally positive and exhibited no nuclear immunoreactivity for beta-catenin. Ten endometrioid carcinomas not exhibiting squamous differentiation were immunoreactive for CDX2; one was focally positive. Electron microscopy in two ovarian endometrioid adenocarcinomas with extensive morular differentiation showed that the morules exhibited epithelial features, but no overt evidence of squamous differentiation. CONCLUSIONS: Typical squamous elements and morules have an overlapping but differing immunophenotype. Morules exhibit no firm immunohistochemical or ultrastructural evidence of squamous differentiation, although immature squamous differentiation cannot be excluded. Nuclear beta-catenin positivity is in keeping with the observation that endometrioid glandular lesions with morules are often associated with beta-catenin gene mutation. The explanation for diffuse nuclear positivity with the intestinal transcription factor CDX2 in morules is not clear, but may be a result of overexpression of nuclear beta-catenin. We suggest that the term morular metaplasia is used instead of squamous morules.

Serous adenocarcinoma of the sigmoid mesentery arising in cystic endosalpingiosis.

This case report describes a Mullerian serous adenocarcinoma arising within a multoloculated cyst lined by ciliated serous-type epithelium located in the sigmoid mesentery. Twenty years previously the patient underwent a hysterectomy, bilateral salpingo-oophorectomy, and omentectomy. The ovaries contained bilateral serous cystadenofibromas, and multiple cysts lined by ciliated serous-type epithelium were present in the omentum. The resection specimen 20 years later contained a 14 cm multiloculated cyst located in the sigmoid mesentery. This was lined largely by benign ciliated serous-type epithelium but a focus of well differentiated serous adenocarcinoma projected into the lumen. Two further peritoneal cysts were present, both of which were lined by ciliated serous-type epithelium. There was a coincidental renal cell carcinoma. This is a unique case of multiple omental, peritoneal, and retroperitoneal cysts (classified as cystic endosalpingiosis), one of which developed a focus of serous adenocarcinoma. Although rarely serous adenocarcinomas, similar to those occurring within the ovary, arise in the retroperitoneum, this is the first reported occurrence in association with a pre-existing benign lesion.

5-HT(2A) receptors stimulate mitogen-activated protein kinase via H(2)O(2) generation in rat renal mesangial cells.

Serotonin (5-HT) stimulates mitogenesis in rat renal mesangial cells through a G protein-coupled 5-HT(2A) receptor. We tested the hypothesis that oxidants might be involved in the signal transduction pathway linking the receptor to extracellular signal-regulated protein kinase (ERK). 5-HT rapidly increased the activity and phosphorylation of ERK. These effects were blocked by the 5-HT(2A) receptor antagonist ketanserin. The peak effect was noted at 5-10 min, and half-maximal stimulation was achieved at 10-30 nM 5-HT. Chemical inhibitor and activator studies supported the involvement of phospholipase C, protein kinase C (PKC), and reactive oxygen species (ROS, i.e., H(2)O(2) and superoxide) generated by an NAD(P)H oxidase-like enzyme in the ERK activation cascade. Mapping studies supported a location for the NAD(P)H oxidase enzyme and the ROS downstream from PKC. Our studies are most consistent with an ERK activation pathway as follows: 5-HT(2A) receptor --> G(q) protein --> phospholipase C --> diacylglycerol --> classical PKC --> NAD(P)H oxidase --> superoxide --> superoxide dismutase --> H(2)O(2) --> mitogen-activated extracellular signal-regulated kinase --> ERK. These studies demonstrate a role for the 5-HT(2A) receptor in rapid, potent, and efficacious activation of ERK in rat renal mesangial cells. They support a role for oxidants in conveying the stimulatory signal from 5-HT, because 1) chemical antioxidants attenuate the 5-HT signal, 2) oxidants and 5-HT selectively activate ERK to a similar degree, 3) 5-HT produces superoxide and H(2)O(2) in these cells, and 4) a specific enzyme [NAD(P)H oxidase] has been implicated as the source of the ROS, which react selectively downstream of classical PKC.

Enrichment of transiently transfected mesangial cells by cell sorting after cotransfection with GFP.

Early passage mesangial cells, like many other nonimmortalized cultured cells, can be difficult to transfect. We devised a simple method to improve the efficiency of transient protein expression under the transcriptional control of promoters in conventional plasmid vectors in rat mesangial cells. We used a vector encoding modified green fluorescent protein (GFP) and sterile fluorescence-activated cell sorting (FACS) to select a population consisting of >90% GFP-expressing cells from passaged nonimmortalized cultures transfected at much lower efficiency. Only 10% transfection efficiency was noted with a beta-galactosidase expression vector alone, but cotransfection with GFP followed by FACS and replating of GFP+ cells yielded greater than fivefold enrichment of cells with detectable beta-galactosidase activity. To demonstrate the expression of a properly oriented and processed membrane protein, we cotransfected GFP with a natriuretic peptide clearance receptor (NPR-C) expression vector. Plasmid-dependent cell surface NPR-C density was enhanced by 89% after FACS, though expression remained lower in selected mesangial cells than in the CHO cell line transfected with the same vector. We conclude that cotransfection of rat mesangial cells with GFP, followed by FACS, results in improvement in transient transfection efficiencies to levels that should suffice for many applications.