Spatiotemporal recapitulation of central nervous system development by murine embryonic stem cell-derived neural stem/progenitor cells

Neural stem/progenitor cells (NS/PCs) can generate a wide variety of neural cells. However, their fates are generally restricted, depending on the time and location of NS/PC origin. Here we demonstrate that we can recapitulate the spatiotemporal regulation of central nervous system (CNS) development in vitro by using a neurosphere-based culture system of embryonic stem (ES) cell-derived NS/PCs. This ES cell-derived neurosphere system enables the efficient derivation of highly neurogenic fibroblast growth factor-responsive NS/PCs with early temporal identities and high cell-fate plasticity. Over repeated passages, these NS/PCs exhibit temporal progression, becoming epidermal growth factor-responsive gliogenic NS/PCs with late temporal identities; this change is accompanied by an alteration in the epigenetic status of the glial fibrillary acidic protein promoter, similar to that observed in the developing brain. Moreover, the rostrocaudal and dorsoventral spatial identities of the NS/PCs can be successfully regulated by sequential administration of several morphogens. These NS/PCs can differentiate into early-born projection neurons, including cholinergic, catecholaminergic, serotonergic, and motor neurons, that exhibit action potentials in vitro. Finally, these NS/PCs differentiate into neurons that form synaptic contacts with host neurons after their transplantation into wild-type and disease model animals. Thus, this culture system can be used to obtain specific neurons from ES cells, is a simple and powerful tool for investigating the underlying mechanisms of CNS development, and is applicable to regenerative treatment for neurological disorders.     

Increased ratio of calcineurin immunoreactive neurons in the caudate nucleus of patients with schizophrenia

Calcineurin (CaN) has been investigated extensively in numerous biochemical, behavioral, and genetic studies in schizophrenia because its function is closely related to dopamine-glutamate signal transduction, which is thought to be associated with pathophysiological changes in schizophrenia. Although evidence has suggested that dysfunction of CaN may be a risk factor for schizophrenia, there have been few reports focusing on the expression of CaN mRNA and CaN protein levels in the brains of schizophrenic patients. In addition, findings on CaN expression in postmortem brains from patients with schizophrenia have been inconsistent. Here, we conducted immunohistochemical examinations of several regions in postmortem brains, including the dorsolateral prefrontal cortex (DLPFC), hippocampus, caudate nucleus, and putamen, using specific antibodies, and compared the results from the brains of nine schizophrenic subjects to nine age- and sex-matched control subjects. There was no significant difference in the ratio of CaN immunoreactive (IR) neurons between schizophrenia and control groups in the DLPFC or hippocampus, and a significantly increased ratio of CaN-IR neurons was seen in the caudate nucleus in the brains from schizophrenia patients. As the striatum contains most of the brain dopamine, the results of the present study have critical implications and suggest that alterations in CaN signaling in the caudate contribute to the pathogenesis of schizophrenia. This is the first report of caudate CaN abnormalities in schizophrenia.     

Shift work and the risk of metabolic syndrome: a nested case-control study

The aim of the present study was to examine the association between shift work and the metabolic syndrome (MetS) using a large-scale longitudinal study design. Data were collected from a historical cohort of health checkups in the Japanese population. The baseline survey, which involved 16,952 inhabitants of the Minami Saku area of the Nagano Prefecture, was started in 1978. A nested case-control study was conducted between 1987 and 1990. This analysis was restricted to 6,712 men and women (age range 25-59 years). A conditional logistic regression model was used to estimate the risk of MetS associated with shift work. Compared with the day workers, shift workers had a significantly higher risk of MetS (odds ratio 1.87; 95% CI, 1.13-3.08). Our results demonstrate that shift work was strongly associated with MetS. The study suggests appropriate dietary habits as a basis for managing the MetS risk of shift workers.     

Anti‐MDA‐5 antibody‐positive clinically amyopathic dermatomyositis with rapidly progressive interstitial lung disease treated with therapeutic plasma exchange: A case series

We present six cases of antimelanoma differentiation‐associated gene 5 antibody (anti‐MDA5‐Ab)‐positive clinically amyopathic dermatomyositis (CADM) with rapidly progressive interstitial lung disease (RP‐ILD), which is known to have a poor prognosis. The outcomes of these cases are described after treatment with therapeutic plasma exchange (TPE). Clinical and therapeutic data for patients with CADM with RP‐ILD were collected retrospectively from medical records. All six patients received early intensive care including high‐dose corticosteroids, intravenous cyclophosphamide, and a calcineurin inhibitor, but lung disease and hypoxia became more severe. TPE was performed over a median of 9.5 sessions (range 3‐14) per patient, and the median duration from admission to TPE was 23 days. Three patients received combined direct hemoperfusion using a polymyxin B‐immobilized fiber column (PMX‐DHP) therapy on successive days to manage acute respiratory failure. Four patients survived and two died due to respiratory failure. In the survival cases, ferritin decreased, and ferritin and KL‐6 were lower at diagnosis. The patients who died had a higher alveolar‐arterial oxygen difference and more severe lung lesions at the time of initiation of TPE. These findings indicate that a combination of conventional therapy and TPE may be useful for improvement of the prognosis of CADM with RP‐ILD at the early stage of onset.     

Changes in programmed death ligand 1 expression in non-small cell lung cancer patients who received anticancer treatments

Background

The expression of programmed death ligand 1 (PD-L1) is considered a predictive biomarker of anti-programmed death 1 (PD-1)/PD-L1 cancer therapies. However, changes in PD-L1 expression of tumor cells during clinical courses have not been fully evaluated. We evaluated changes in PD-L1 expression for non-small cell lung cancer (NSCLC) patients who received anticancer treatments during clinical courses.

Methods

In 76 NSCLC patients, PD-L1 expression was evaluated before and after anticancer treatment by immunohistochemical (IHC) analysis using an anti-PD-L1 antibody. We defined two cut-off points of PD-L1 expression (1 and 50%) and three corresponding IHC groups (A: 0%, B: 1–49%, and C: ≥50%). IHC group B and C were considered to be positive expression, and we defined the difference of IHC group between pre- and post-treatment as ‘major change’ in PD-L1 expression.

Results

Before anticancer treatment, PD-L1 expression was observed in 38/76 (50%) patients, and was significantly less common in patients harboring mutations in the epidermal growth factor receptor gene (EGFR) than in those without (P?=?0.039). After anticancer treatment, PD-L1 expression was observed in 36/76 (47%) patients. Major increases in PD-L1 expression were seen in 11 (14%), and major decreases in 18 (24%) patients. Among 13 patients harboring EGFR mutations treated with EGFR tyrosine-kinase inhibitor (EGFR-TKI), five (38%) showed major increases.

Conclusion

Major changes of PD-L1 expression in tumor cells were observed in 38% of NSCLC patients who received anticancer treatments. And, treatments with EGFR-TKI may increase PD-L1 expression in NSCLC patients harboring EGFR mutations.

     

Highly sensitive detection of <Emphasis Type="Italic">ESR1</Emphasis> mutations in cell-free DNA from patients with metastatic breast cancer using molecular barcode sequencing

Purpose

We aimed to develop a highly sensitive method to detect ESR1 mutations in cell-free DNA (cfDNA) using next-generation sequencing with molecular barcode (MB–NGS) targeting the hotspot segment (c.1600–1713).

Methods

The sensitivity of MB–NGS was tested using serially diluted ESR1 mutant DNA and then cfDNA samples from 34 patients with metastatic breast cancer were analyzed with MB–NGS. The results of MB–NGS were validated in comparison with conventional NGS and droplet digital PCR (ddPCR).

Results

MB–NGS showed a higher sensitivity (0.1%) than NGS without barcode (1%) by reducing background errors. Of the cfDNA samples from 34 patients with metastatic breast cancer, NGS without barcode revealed seven mutations in six patients (17.6%) and MB–NGS revealed six additional mutations including three mutations not reported in the COSMIC database of breast cancer, resulting in total 13 ESR1 mutations in ten patients (29.4%). Regarding the three hotspot mutations, all the patients with mutations detected by MB–NGS had identical mutations detected by droplet digital PCR (ddPCR), and mutant allele frequency correlated very well between both (r = 0.850, p < 0.01). Moreover, all the patients without these mutations by MB–NGS were found to have no mutations by ddPCR.

Conclusion

In conclusion, MB–NGS could successfully detect ESR1 mutations in cfDNA with a higher sensitivity of 0.1% than conventional NGS and was considered as clinically useful as ddPCR.

     

Expression of gremlin1 in gastric cancer and its clinical significance

As an antagonist of bone morphogenetic proteins (BMPs), 2, 4 and 7, gremlin1 plays a role in regulating organogenesis, tissue differentiation and angiogenesis. However, there is little information regarding gremlin1 in gastrointestinal cancer. We attempted to clarify how gremlin1 expression affects the clinical features and biological properties of gastric cancer. A total of 232 gastric cancer patients who received R0 gastrectomy at Kagoshima University Hospital were enrolled. Gremlin1 expression in gastric cancer was detected by immunohistochemical and western blotting methods. Correlations between clinicopathological parameters and gremlin1 expression were analyzed. Gremlin1 was identified in the cytoplasm and nucleus of all gastric cancer cell lines and some regions of surgical specimens of gastric cancer. One hundred and seventeen of the 232 patients (50.4%) were classified as gremlin1-positive based on gremlin1 expression. Gremlin1 positivity was correlated with shallower tumor depth, smaller tumor size, less nodal involvement and vessel invasion (p < 0.05). The 5-year survival rate of the gremlin1-positive group was 81%, which was significantly higher than the gremlin1-negative group (p < 0.01). Multivariate analysis revealed that gremlin1 was not selected as an independent prognostic marker. Gremlin1 expression in gastric cancer may be a useful prognostic marker that is involved with the BMP signaling pathway. Furthermore, gremlin1 may have clinical use as a diagnostic and treatment tool.     

THE ROLE OF ENDOTHELIAL CELLS IN THE PATHOGENESIS OF ATHEROSCLEROSIS

Using mainly rabbits and rats investigations on the endothelial permeability of the aorta, on relation between thrombus formation and endothelial damage, and on tissue culture of the endothelial cells of the rabbit aorta have been carried out.
The permeability of endothelial layer of rabbit aorta Increased by cholesterol-feeding. This seems to be derived from morphological and functional disorder of the endothelial cells. As a hemodynamic effect of blood flow, increased permeability of the endothelium of rat aorta was also found in experimental renal hypertension.
Accumulation of blood-plasma components (edema) was seen in the subendothellal area in cholesterol-fed rabbit aorta and in hypertensive rat aorta. Long-standing subendothellal edema is considered to be the cause of fibrous intlmal thickening (Grade I lesion/Nakashlma).
Endothelial defect of the aorta due to various damages is related to thrombus formation, and this seems to be accelerated by cholesterol feeding.
Morphological differences between normal endothelium and that of atherosclerotic lesion in human and in rabbit aortas can be seen.
Tissue culture of the endothelium was performed in vivo and in vitro, and some discussions were made on the characteristics of cultured endothelial cells.