Multiple system atrophy (MSA) is a fatal adult-onset neurodegenerative disease that is characterized by varying degrees of cerebellar dysfunction and Parkinsonism. The neuropathological hallmark of MSA is alpha-synuclein (AS)-positive glial cytoplasmic inclusions (GCIs). Although severe neuronal loss (NL) is also observed in MSA, neuronal inclusions (NIs) are rare compared to GCIs, such that the pathological mechanism of NL in MSA is unclear. GCIs and NIs are late-stage pathology features relative to AS oligomers and may not represent early pathological changes in MSA. To reveal the early pathology of MSA, it is necessary to examine the early aggregation of AS, i.e., AS oligomers. Here, we adopted a proximity ligation assay (PLA) to examine the distribution of AS oligomers in brain tissue samples from patients with MSA and other diseases. Surprisingly, MSA brains showed a widespread distribution and abundant accumulation of oligomeric AS in neurons as well as oligodendrocytes of the neocortex. In several regions, oligomeric AS signal intensity was higher in cases with MSA than in cases with Parkinson's disease. In contrast to previous studies, AS-PLA revealed abundant AS oligomer accumulation in Purkinje cells in MSA brains, identifying oligomeric AS accumulation as a possible cause of Purkinje cell loss. This wide distribution of AS oligomers in MSA brain neurons has not been described previously and indicates a pathological mechanism of NL in MSA.

The pathological hallmarks of Parkinson’s disease (PD) are α-synuclein (αSYN)-positive inclusions referred to as Lewy bodies and Lewy neurites, collectively referred to as Lewy-related pathology (LRP). LRP is thought to propagate in an ascending manner throughout the brain as the disease progresses. LRP is visible with histologic methods and is thought to represent a later stage of the disease process, while αSYN oligomers, which are not visible with routine histologic methods, are considered earlier. There is increasing evidence to suggest that αSYN oligomers may be more toxic than visible LRP. Detecting αSYN oligomers requires special techniques, and their distribution and association with clinical features are important research objectives. In this report, we describe the distribution of αSYN oligomers in multiple cortical and subcortical regions of PD using a proximity ligation assay (PLA). We observe widespread distribution of αSYN oligomers with PLA and more restricted distribution of LRP with αSYN immunohistochemistry. The distribution of αSYN oligomers differed from LRP in that αSYN oligomer burden was significantly greater in the neocortex, while LRP was greater in vulnerable subcortical regions, including the brainstem. We also found that cognitive impairment was associated with αSYN oligomers in the hippocampus. These results suggest that αSYN oligomers may be widely distributed in PD early in the disease process and that they may contribute to cognitive impairment in PD.

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Although various methods for collagen gel-based cell invasion assays have been described, there continues to be a need for a simpler and more objective assay. Here, we describe an easy-to-prepare double-layered collagen gel hemisphere (DL-CGH) system that satisfies these requirements, and we demonstrate the advantages of this new system for visualizing cell movements during invasion. DL-CGH consists of a central core collagen layer surrounded by an outer cover collagen layer. A droplet of collagen I solution (containing cells to be examined) naturally forms a small hemisphere on the bottom of the culture dish. After this central core layer gels, a second droplet is placed atop the first gel, encapsulating it completely. The hemisphere is submerged in the medium and cultured. The invasive activity of cells that infiltrate from the inner to the outer layer can be evaluated optically. Using this in vitro system, we measured the inhibitory effect of E-cadherin expression on cancer cell invasion. DL-CGH also allowed visualization of interactions between invading cancer cells and the stroma. Cancer cells, which lack the proteases required for direct entrance into the three-dimensional collagen matrix, were seen to slip like amoebas through matrix gaps generated by the pericellular proteolytic activity of fibroblasts. [Supplementary materials are available for this article. Go to the publisher's online edition of Cell Communication and Adhesion for the following free supplemental resources: Movies 1-3; 4a and b].

Abstract. In this study, chemosensitivity tests were performed on both primary lesions (Pls) and lymph node metastases (lMs) from surgically resected non-small cell lung cancer (nsclc). Differences between the results obtained were evaluated. operative specimens were obtained from 13 patients with nsclc [6 with squamous cell carcinoma (sQ) and 7 with adenocarcinoma (AD)] whose lymph nodes were confirmed to be positive for metastasis. Both the Pl and lM from the same patient were examined immediately after resection. the collagen gel droplet-embedded culture drug sensitivity test (cD-Dst) was used as the chemosensitivity test against six anticancer drugs [5-fluorouracil (5-FU), cisplatin, gemcitabine, docetaxel, vinorelbine and sn-38 (an active metabolite of irinotecan)]. When the growth rate, determined by the t/c ratio (t, signal for viable cells in the treated group and c, signal in the control) was less than 50%, the tumor cells were considered to be sensitive to the drug. only in 4 cases (2 sQ and 2 AD) was the chemosensitivity of the primary lesion identical to that of lM. In the sQ cases, chemosensitivity of the primary lesions to 5-FU tended to be consistent with that of LMs. In contrast, the primary lesions in 4 of the 7 AD cases were negative for chemosensitivity to 5-FU; however, LMs were sensitive. In many cases, the chemosensitivity of the Pls to each anticancer drug differed from that of the lMs. In conclusion, both primary and metastatic tumors should be examined to ensure maximum clinical efficacy of in vitro drug-sensitivity testing for adjuvant chemotherapy after complete resection of n1 and n2 nsclc.Introduction the effectiveness of chemotherapy directly affects the prognosis of advanced lung cancer. Adjuvant chemotherapy is expected to contribute to the improvement of the prognosis of patients who undergo curative surgical treatments. the clinical outcome of chemotherapy against non-small cell lung cancer (nsclc) has improved since new drugs have been developed and their combination therapies have been established. However, these drugs are not always equally efficacious in all NSCLC patients. A drug that has curative effects on some patients has sometimes been observed to cause only side effects in other patients. These findings suggest the need for the appropriate selection of drugs separately for each individual patient. certain in vitro chemosensitivity tests have been established for the identification of an effective anticancer drug for individual cases (1-3), and the efficacy of these assays has been described in many clinical reports on lung cancer (4-9) and other malignancies (10-15).Anticancer drugs are expected to be effective not only against primary but also against metastatic lesions. Specifically, adjuvant chemotherapy, after complete resection of lung cancer, targets putative dormant metastasis. However, in certain clinical cases the chemosensitivity of metastatic lesions differs from that of primary lesions. this appears to be normal since the pathological features of the prima...