Intratumoural immune cell landscape in germinoma reveals multipotent lineages and exhibits prognostic significance.

AIMS: Alterations in microenvironments are a hallmark of cancer, and these alterations in germinomas are of particular significance. Germinoma, the most common subtype of central nervous system germ cell tumours, often exhibits massive immune cell infiltration intermingled with tumour cells. The role of these immune cells in germinoma, however, remains unknown. METHODS: We investigated the cellular constituents of immune microenvironments and their clinical impacts on prognosis in 100 germinoma cases. RESULTS: Patients with germinomas lower in tumour cell content (i.e. higher immune cell infiltration) had a significantly longer progression-free survival time than those with higher tumour cell contents (P = 0.03). Transcriptome analyses and RNA in-situ hybridization indicated that infiltrating immune cells comprised a wide variety of cell types, including lymphocytes and myelocyte-lineage cells. High expression of CD4 was significantly associated with good prognosis, whereas elevated nitric oxide synthase 2 was associated with poor prognosis. PD1 (PDCD1) was expressed by immune cells present in most germinomas (93.8%), and PD-L1 (CD274) expression was found in tumour cells in the majority of germinomas examined (73.5%). CONCLUSIONS: The collective data strongly suggest that infiltrating immune cells play an important role in predicting treatment response. Further investigation should lead to additional categorization of germinoma to safely reduce treatment intensity depending on tumour/immune cell balance and to develop possible future immunotherapies.

Integrin antagonist augments the therapeutic effect of adenovirus-mediated REIC/Dkk-3 gene therapy for malignant glioma.

Reduced expression in immortalized cells/Dickkopf-3 (REIC/Dkk-3) was identified as a gene whose expression is reduced in many human cancers. REIC/Dkk-3 expression is also downregulated in malignant glioma and regulates cell growth through caspase-dependent apoptosis. cRGD (EMD121974), an antagonist of integrins, has demonstrated preclinical efficacy against malignant glioma. In this study, we investigated the antiglioma effect of combination therapy using an adenovirus vector carrying REIC/Dkk-3 (Ad-REIC) and cRGD. Quantitative real-time reverse-transcription PCR revealed the reduction of REIC/Dkk-3 mRNA levels in malignant glioma cell lines. The reduction of REIC/Dkk-3 protein expression in malignant glioma cell lines was also confirmed with western blot analysis. After treatment with Ad-REIC and cRGD, the proliferative rate of malignant glioma cells was significantly reduced in a time-dependent manner. In vivo, there was a statistically significant increase in the survival of mice treated with Ad-REIC and cRGD combination therapy compared with Ad-REIC monotherapy. We identified an apoptotic effect following monotherapy with Ad-REIC. Moreover, cRGD augmented the antiglioma efficacy of Ad-REIC. These results may lead to a promising new approach for the treatment of malignant glioma.

Detailed Arterial Anatomy and Its Anastomoses of the Sphenoid Ridge and Olfactory Groove Meningiomas with Special Reference to the Recurrent Branches from the Ophthalmic Artery.

BACKGROUND AND PURPOSE: Detailed arterial anatomy of the sphenoid ridge and olfactory groove meningiomas is complicated due to the fine angioarchitecture and anastomoses between each feeder. Herein, we present details of the arterial anatomy and the relationships of feeders in these lesions. MATERIALS AND METHODS: This study included 20 patients admitted to our department between April 2015 and March 2020. Conditions of subjects consisted of 16 sphenoid ridge meningiomas and 4 olfactory groove meningiomas. We mainly analyzed arterial anatomy using 3D rotational angiography and slab MIP images of these lesions. We also analyzed the anastomoses of each feeder. RESULTS: We found that 19 (95%), 15 (75%), and 15 (75%) lesions had feeders from the ophthalmic, internal carotid, and external carotid arteries, respectively. As feeders from the ophthalmic artery, recurrent meningeal arteries were involved in 18 lesions (90%). Fifteen lesions (75%) had anastomoses between each feeder. CONCLUSIONS: Most of the meningiomas in the sphenoid ridge and olfactory groove had feeders from the ophthalmic and internal carotid arteries. There were various anastomoses between each feeder. This is the first report to demonstrate the detailed arterial anatomy and frequency of recurrent branches from the ophthalmic artery and their anastomoses using detailed imaging techniques.

Fibroblast growth factor 13 regulates glioma cell invasion and is important for bevacizumab-induced glioma invasion.

Glioblastoma has the poorest prognosis, and is characterized by excessive invasion and angiogenesis. To determine the invasive mechanisms, we previously used two glioma cell lines (J3T-1 and J3T-2) with different invasive phenotypes. The J3T-1 showed abundant angiogenesis and tumor cell invasion around neovasculature, while J3T-2 showed diffuse cell infiltration into surrounding healthy parenchyma. Microarray analyses were used to identify invasion-related genes in J3T-2 cells, and the expressed genes and their intracellular and intratumoral distribution patterns were evaluated in J3T-2 cell lines, human glioma cell lines, human glioblastoma stem cells and human glioblastoma specimens. To determine the role of the invasion-related genes, invasive activities were evaluated in vitro and in vivo. Fibroblast growth factor 13 (FGF13) was overexpressed in J3T-2 cells compared to J3T-1 cells, and in human glioma cell lines, human glioblastoma stem cells and human glioblastoma specimens, when compared to that of normal human astrocytes. Immunohistochemical staining and the RNA-seq (sequencing) data from the IVY Glioblastoma Atlas Project showed FGF13 expression in glioma cells in the invasive edges of tumor specimens. Also, the intracellular distribution was mainly in the cytoplasm of tumor cells and colocalized with tubulin. Overexpression of FGF13 stabilized tubulin dynamics in vitro and knockdown of FGF13 decreased glioma invasion both in vitro and in vivo and prolonged overall survival of several xenograft models. FGF13 was negatively regulated by hypoxic condition. Silencing of FGF13 also decreased in vivo bevacizumab-induced glioma invasion. In conclusion, FGF13 regulated glioma cell invasion and bevacizumab-induced glioma invasion, and could be a novel target for glioma treatment.

Importance of lymph vessels in gastric cancer: a prognostic indicator in general and a predictor for lymph node metastasis in early stage cancer.

BACKGROUND: Metastasis to regional lymph nodes (LNs) through lymphatic vessels is common in cancer progression and is an important prognostic factor in many cancers. Recent evidence suggests that tumour lymphangiogenesis promotes lymphatic metastasis. AIMS: To study the role of lymph vessel density (LVD) in gastric cancer and investigate whether LVD is associated with LN metastasis/prognosis. METHODS: Lymphatics of 117 primary human gastric cancer cases were investigated by quantitative immunohistochemical staining for podoplanin. The relation between LVD and LN metastasis and other established clinicopathological parameters was analysed. The relation between LVD and prognosis was also studied. RESULTS: Mean LVD of "hot spots" was 11.6/case. LVD significantly correlated with LN and podoplanin positive lymphatic invasion. High LVD was associated with worse overall survival. In multivariate analysis, positive LVD was a significant independent predictor of overall survival, depth of invasion, and TNM stage. LVD significantly correlated with LN metastasis at surgery and podoplanin positive lymphatic invasion. In multivariate analysis, positive LVD was an independent significant predictor of LN metastasis. CONCLUSIONS: Increased podoplanin expression is significantly associated with LN metastasis, and may play an important role in detecting LN metastasis in gastric cancer. Furthermore, LVD may be a significant prognostic factor in gastric cancer at any stage. In addition, LVD and lymph vessel invasion detected by podoplanin immunohistochemistry are associated with LN metastasis in T1 early gastric cancer. LVD assessment by podoplanin immunohistochemistry may become a useful predictor of LN metastasis in T1 early gastric cancer and may influence the decision making process for additional surgery.

Generation of inositol phosphates, cytosolic Ca2+, and secretion of noradrenaline in PC12 cells treated with glutamate.

Glutamate transiently stimulated rat pheochromocytoma PC12 cells and caused an inositol trisphosphate formation and an increase in levels of Ca+ in the cytosol. The rank order of potency of glutamate greater than N-methyl-D-aspartate (NMDA) much greater than kainate = quisqualate is characteristic of an interaction with NMDA receptors. The effect of glutamate on inositol trisphosphate formation disappeared in a low Mg2+ buffer and was not blocked by DL-2-amino-5-phosphonovalerate, an antagonist for NMDA receptors coupled to ion channels. Although glutamate failed to stimulate noradrenaline secretion, glutamate enhanced the effect of bradykinin, but not of Ca ionophore A23187, or KCl. These results suggest the existence of metabotropic glutamate receptors, different from previously reported receptors, in PC12 cells.

Sex-related difference in oxidative metabolism of testosterone and erythromycin by hamster liver microsomes.

The activities of testosterone hydroxylases and erythromycin N-demethylase were significantly higher in liver microsomes from female hamsters than in the male counterparts. SDS-polyacrylamide gel electrophoresis revealed a difference in protein composition between male and female liver microsomes in the molecular mass region comprising cytochrome P-450. Western blot analysis showed further that antibodies to rat male-specific cytochrome P-450 crossreacted with at least two proteins in both male and female hamster microsomes, but one of the female proteins had a different molecular mass from that of the male proteins. It is concluded that sex difference in liver microsomal cytochrome P-450 is not restricted to rats and mice, as has previously been believed.

Experience with the high occipital transcortical approach in the treatment of intraventricular hemorrhage. Report of two cases.

Two patients with intraventricular hemorrhage (IVH) were treated by direct removal of their intraventricular hematomas via a high occipital transcortical approach with successful results. This approach lies between the parietooccipital transcortical approach and the occipital transcortical approach. The patients were a 90-year-old woman with idiopathic IVH and a 60-year-old man with hemorrhage caused by bleeding in the thalamus. In both cases, the hematoma was tightly packed in the lateral ventricle. In the former case, the inferior horn of the lateral ventricle was extremely swollen, and the patient was at risk for development of uncal herniation. With the goals of complete elimination of the hematoma in the inferior horn and identification of the source of bleeding, a high occipital transcortical approach was applied, and the hematoma was removed under direct vision. With the patient in the lateral position, a minor craniotomy of approximately 3 cm was performed around the puncture site of the posterior horn (8 cm craniad from the inion and 3 cm lateral from the midline). A 1-cm cortical incision was made and the posterior horn was reached. First, the portion of hematoma at this site was removed, and then the remainder was completely removed from the interior horn and corpus. Using this method, the entire region of the lateral ventricle, including the inferior horn, corpus, and posterior horn, can be covered in a single operative field, and it is also possible to have sufficient working space for the operation.

[Two cases of severe ischemic mitral regurgitation treated with CABG alone].

We report here two cases in which patients fell into pulmonary edema due to ischemic mitral regurgitation (ischemic MR) after cardiac catheterization and underwent emergency coronary artery bypass grafting (CABG) using an intra-aortic balloon pumping. The patient were a 65-year-old man and a 80-year-old woman, and both had a chief complaint of angina after myocardiac infarction. In both cases, coronary angiography revealed triple vessel disease, and left ventriculography showed severe MR. However echocardiography, when they were hospitalized, did not show significant MR. Therefore we thought that they had gone into congestive heart failure because cardiac ischemia and volume load following cardiac catheterization provoked MR. In fact, postoperative left ventriculography and echocardiography showed decreased MR. We now think that it is important to keep in mind the cases of severe ischemic MR for which CABG alone is adequate treatment and to evaluate ischemic MR not only by left ventriculography but also by echocardiography.

[Single-stage management of postoperative sternal wound infection using a pectoral musculocutaneous flap].

Two patients with postoperative sternal wound infection were successfully treated by a pectral musculocutaneous flap. A single-stage procedure of debridement and immediate closure with a pectral musculocutaneous flap can eliminate irrigation, open wound management, or reoperation for closure. Therefore, this method is very safe, simple, and effective for the management of sternal wound infections.

[Experience of double internal thoracic artery grafts in coronary artery revascularization].

We have started employing two internal thoracic artery grafts in coronary artery revascularization since February 1988. We have carried out seven such operative procedures in a 15 month period since then. Ages ranged from 55 to 67 years with a mean of 60.6. Six patients were male and one patient was female. One patient was in Canadian Cardiovascular Society angina Class I, one in Class II, four in Class III, and one in Class IV, preoperatively. There were no operative deaths. Postoperatively, six patients were in CCS Class I and one was in Class II. All of the internal thoracic artery grafts were patent at hospital discharge in six angiographically examined patients. We feel that use of double internal thoracic artery grafts in coronary revascularization carries minimal additional risk, when performed in selected case.

The integrin inhibitor cilengitide enhances the anti-glioma efficacy of vasculostatin-expressing oncolytic virus.

Oncolytic viral (OV) therapy has been considered as a promising treatment modality for brain tumors. Vasculostatin, the fragment of brain-specific angiogenesis inhibitor-1, shows anti-angiogenic activity against malignant gliomas. Previously, a vasculostatin-expressing oncolytic herpes simplex virus-1, Rapid Antiangiogenesis Mediated By Oncolytic virus (RAMBO), was reported to have a potent antitumor effect. Here, we investigated the therapeutic efficacy of RAMBO and cilengitide, an integrin inhibitor, combination therapy for malignant glioma. In vitro, tube formation was significantly decreased in RAMBO and cilengitide combination treatment compared with RAMBO or cilengitide monotherapy. Moreover, combination treatment induced a synergistic suppressive effect on endothelial cell migration compared with the control virus. RAMBO, combined with cilengitide, induced synergistic cytotoxicity on glioma cells. In the caspase-8 and -9 assays, the relative absorption of U87ΔEGFR cell clusters treated with cilengitide and with RAMBO was significantly higher than that of those treated with control. In addition, the activity of caspase 3/7 was significantly increased with combination therapy. In vivo, there was a significant increase in the survival of mice treated with combination therapy compared with RAMBO or cilengitide monotherapy. These results indicate that cilengitide enhanced vasculostatin-expressing OV therapy for malignant glioma and provide a rationale for designing future clinical trials combining these two agents.

Therapeutic effect of suicide gene-transferred mesenchymal stem cells in a rat model of glioma.

We evaluated a new therapeutic strategy for malignant glioma, which combines intratumoral inoculation of mesenchymal stem cells (MSCs) expressing cytosine deaminase gene with 5-fluorocytosine (5-FC) administration. For in vitro and in vivo experiments, MSCs were transfected with adenovirus carrying either enhanced green fluorescent protein gene (AdexCAEGFP) or cytosine deaminase gene (AdexCACD), to establish MSC-expressing EGFP (MSC-EGFP) or CD (MSC-CD). Co-culture of 9L glioma cells with MSC-CD in a medium containing 5-FC resulted in a remarkable reduction in 9L cell viability. The migratory ability of MSC-EGFP toward 9L cells was demonstrated by double-chamber assay. For the in vivo study, rats harboring 9L brain tumors were inoculated with MSC-EGFP or MSC-CD. Immunohistochemistry of rat brain tumors inoculated with MSC-EGFP showed intratumoral distribution of MSC-EGFP. Survival analysis of rats bearing 9L gliomas treated with intratumoral MSC-CD and intraperitoneal 5-FC resulted in significant prolongation of survival compared with control animals. In conclusion, molecular therapy combining suicide gene therapy and MSCs as a targeting vehicle represents a potential new therapeutic approach for malignant glioma, both with respect to the antitumor potential of this system and its neuroprotective effect on normal brain tissue.