Morphometric investigation of the hippocampal formation in Ihara's genetically epileptic rats using magnetic resonance images and histopathology

The newly developed Ihara's genetically epileptic rat (IGER) is thought to be a useful tool for studying epilepsy of the limbic type. Morphological changes in the hip-pocampus, which is one of the characteristic findings in the brains of patients with temporal lobe epilepsy, were presumed to occur also in IGER during the course of epileptic activities. We investigated chronological changes in volume of the IGER hippocampal formation using mag-netic resonance (MR) imaging. The experiment consisted of two protocols. First, MR imaging was performed for volumetric measurement of the cerebrum and the hippocampus in four male IGER, each at 2, 4, 6, 8, 10 and 12 months of age. After MR examination, the brains were subjected to histological and morphometric examinations. Second, changes in the hippocampal volume were traced chronologically in three IGER at the ages of 2, 4, 6, 8 and 10 months. Age-matched Wistar rats were used as controls. In IGER, the volume of the cerebrum, especially of the hippocampus, and the ratio of the hippocampal to the cerebral volume increased, in concert with the increase in seizure activities. Histological and morphometric invest-igations revealed that there was mild astrogliosis and marked microgliosis with hypertrophic change in the hippocampal formation with aging, but no loss of neurons was observed. Neither an increase in volume nor gliosis was found in the brains of control rats. The enlargement of the hippocampal formation in IGER brains was assumed to be partially due to gliosis of astrocyte and microglia with hypertrophy.     

A case of glioblastoma multiforme which indicated the early stage on brain MRI]

A 57-year-old male was urgently carried to our hospital because of sudden loss of consciousness, lasting about 10 minutes. He had resumed consciousness before he arrived at our hospital. Neurologically, he had mild muscle weakness of the right arm. Deep tendon reflexes in the right upper extremity were reduced. In high level functions, speech disturbance, dysgraphia (disturbed ability to write Hiragana), and constructive apraxia were noted. A brain MRI upon admission showed a poorly demarcated, high signal intensity area in the cortical and subcortical layers of the left temporal and parietal lobes. This was visible on T 2 weighted images(T 2 WI), although no abnormalities were visible on T 1 weighted images(T 1 WI). No contrast enhancement was effected by Gd-DTPA. The patient was therefore suspected of having a tumor or degenerative disease and was monitored closely. About 4 months later after onset, his symptoms became aggravated, and brain MRI disclosed a marked low signal intensity area on T 1 WI and a heterogeneous high signal intensity area on T 2 WI. The abnormal signal intensity area was surrounded by extensive edema and mass effect. Ring-shaped, irregular, contrast enhanced areas were also visible. Cerebral angiography revealed a poorly demarcated tumor stain in the area supplied by the middle cerebral artery. The tumor was removed surgically and was histopathologically rated as glioblastoma multiforme(GBM). Because this case represents a valuable example of early stage of GBM, it will be discussed in this paper, along with differential diagnoses.     

In vitro study on the removal of pemphigus antibody with immunoadsorption

The adsorbability of pemphigus vulgaris (PV) antibodies to immunoadsorbent with phenylalanine, tryptophan, and dextran sulfate as ligands was examined, PV antibodies were adsorbed to immunoadsorbents with both phenylalanine and tryptophan as ligands, in batchwise adsorption and column adsorption. PV antibodies were not adsorbed to immunoadsorbent with dextran sulfate as ligand, in column adsorption. It was also shown that the rates of decrease of normal protein fractions in column adsorption with phenylalanine as a ligand were small. Our results suggest that immunoadsorption therapy using sorbent with phenylalanine as a ligand may be useful for the treatment of PV. © 1996 Wiley-Liss, Inc.     

The effectiveness of new triple combination therapy using synthetic disease-modifying anti-rheumatic drugs with different pharmacological function against rheumatoid arthritis: the verification by an in vitro and clinical study

The study aims to confirm the feasibility of new oral triple combination therapy using methotrexate (MTX), mizoribine (MZR), and tacrolimus (TAC) in patients with rheumatoid arthritis (RA) by in vitro and clinical analyses. Triple therapy with a combination of MTX, MZR, and TAC was used for an in vitro study with osteoclasts and a prospective clinical study in order to show the efficacy of these agents against refractory RA. In particular, low-dose TAC or MZR was added to treat 14 patients with RA that was resistant to MTX + MZR or MTX + TAC dual therapy. The combination of three pharmacological agents showed statistically significant differences to reduce differentiation induction and activity of osteoclasts compared with single and double agents. In clinical use, triple therapy showed a statistically significant difference in the improvement of Disease Activity Score-28-erythrocyte sedimentation rate and the Simple Disease Activity Index score at around 8 months. Additionally, the serum matrix metalloproteinase-3 level significantly decreased. No patients dropped out because of adverse effects. Based on this in vitro and prospective clinical study, oral triple therapy might be effective against refractory RA. Furthermore, this therapy might be safe and economical for clinical practice.     

Autoimmune neutropenia due to antineutrophil antibodies in a patient with primary sclerosing cholangitis

Autoimmune neutropenia (AIN) is defined as a decrease in the circulating absolute neutrophil count (ANC) to less than 1500/μl caused by serum antineutrophil antibodies. Secondary AIN is associated with various autoimmune diseases. Herein we present the case of a patient with primary sclerosing cholangitis (PSC) who developed secondary AIN. A 19-year-old man was admitted due to liver injury, and a diagnosis of PSC was established by cholangiogram and liver biopsy. Severe neutropenia, with the ANC down to 130/μl, developed during his hospital course. No medications had been given at that time and bone marrow aspiration revealed no abnormality. Therefore we suspected secondary AIN as a causative etiology and examined whether antineutrophil antibodies were detectable in the patient’s sera by flow cytometric analysis of the granulocyte indirect immunofluorescence test. We found that antineutrophil antibody was strongly positive on admission, and the titer decreased along with recovery from neutropenia. This is the first reported case of a patient with PSC who developed AIN, with detection of serum antineutrophil antibodies.     

Human T-cell leukemia virus type-I (HTLV-I)-specific T-cell responses detected using three-divided glutathione-S-transferase (GST)-Tax fusion proteins

Insufficient T-cell response to human T-cell leukemia virus type-I (HTLV-I) is a potential risk factor in adult T-cell leukemia (ATL). We established an assay system for detecting HTLV-I-specific T-cell response by using recombinant glutathione-S-transferase (GST) proteins fused with HTLV-I Tax protein that was divided into three portions, Tax-A, -B, and -C, corresponding to the N-terminal, central and C-terminal regions, respectively. When splenocytes from rats immunized with plasmids encoding Tax cDNA were incubated with these recombinant proteins, strong interferon gamma (IFN-gamma-producing responses occurred against GST-Tax proteins but not against control GST proteins. No such Tax-specific responses were observed in splenocytes from naive rats. Cocktails of oligopeptides corresponding to the Tax-A, -B, and -C regions also induced IFN-gamma-producing responses when incubated with splenocytes from immunized rats, but required higher amounts of antigens and there were a shorter periods of sustained T-cell responses than with GST-Tax protein-based assay. Although splenocytes from immunized rats predominantly reacted against GST-Tax-B protein, they failed to react with peptide cocktails corresponding to the Tax-B region, likely because the major epitope was interrupted in the initially prepared series of peptides. Using a newly prepared peptide series we found that splenocytes predominantly reacted with a peptide located in the Tax-B region that overlaps with a previously identified cytotoxic T lymphocytes (CTL) epitope of this rat strain. Using this system, we examined peripheral blood mononuclear cells (PBMC) from an ATL patient who underwent complete remission following hematopoietic stem cell transplantation (HSCT). PBMC from this patient produced a significant Tax-specific T-cell response predominantly against GST-Tax-A protein. This is consistent with the previous finding that this patient exhibited a strong HLA-A2-restricted CTL response to Tax 11-19 epitope, which is located in the Tax-A region. This study provides a diagnostic tool, useful for monitoring HTLV-I-specific T-cell immunity in patients and for surveying HTLV-I-carriers to identify an immunological group at high risk for ATL development, regardless of their human leukocyte antigen (HLA) types. It is also useful for predicting the location of T-cell epitopes, which may be applicable in future vaccine strategies.     

Identification of CXCL5/ENA-78 as a factor involved in the interaction between cholangiocarcinoma cells and cancer-associated fibroblasts

Knowledge of tumor-stromal interactions is essential for understanding tumor development. We focused on the interaction between cholangiocarcinoma and cancer-associated fibroblasts (CAFs) in intrahepatic cholangiocarcinoma and reported their positive interaction in vitro and in vivo. The aim of this study is to identify the key protein involved in the interaction between cholangiocarcinoma cells and CAFs and its role on cholangiocarcinoma progression. Using the conditioning medium from cholangiocarcinoma cells, hepatic stellate cells and coculture of them, Protein-Chip analysis with SELDI-TOF-MS showed that the peak of an 8,360-Da protein remarkably increased in the coculture medium. This protein was identified as CXCL5/ENA78, epithelial cell-derived neutrophil-activating peptide-78, by q-TOF/MS/MS analysis. Two cholangiocarcinoma cell lines, HuCCT1 and RBE, produced CXCL5 that promoted their invasion and migration in an autocrine fashion. These effects of CXCL5 significantly decreased by inhibition of CXC-receptor 2, which is the receptor for CXCL5. In addition, IL-1β produced by hepatic stellate cells induced the expression of CXCL5 in cholangiocarcinoma cells. In human tissue samples, a significant correlation was observed between CAFs and CXCL5 produced by cholangiocarcinoma cells in intrahepatic cholangiocarcinoma (p = 0.0044). Furthermore, the high-CXCL5-expression group exhibited poor overall survival after curative hepatic resection (p = 0.027). The presence of tumor-infiltrating neutrophils expressing CD66b was associated with CXCL5 expression in tumor cells (p < 0.0001). These data suggest that CXCL5 is important for the interaction between cholangiocarcinoma and CAFs, and inhibition of tumor-stromal interactions may be a useful therapeutic approach for cholangiocarcinoma.     

Personalized peptide vaccination in patients with refractory non-small cell lung cancer

Since the prognosis of non-small cell lung cancer (NSCLC) remains poor, the development of novel therapeutic approaches, including cancer vaccines, is highly desirable. In the current study, we conducted a phase II study of personalized peptide vaccination (PPV), in which a maximum of 4 peptides were selected based on pre-existing humoral immune responses and administered subcutaneously (weekly for 6 consecutive weeks and bi-weekly thereafter) in refractory NSCLC patients. Forty-one refractory NSCLC patients (4 stage IIIb, 22 stage IV and 15 recurrent), who had failed to respond to chemotherapy and/or targeted therapy (median number of regimens, 3; median duration, 10 months), were enrolled. Median overall survival (OS) was 304 days with a one-year survival rate of 42% in the enrolled patients. The main toxicity of PPV was skin reactions at the injection sites, but no serious adverse events were observed. In order to identify potential biomarkers for predicting OS, pre-vaccination and post-vaccination clinical findings and laboratory data were retrospectively assessed and evaluated by multivariate Cox regression analysis. Among the pre-vaccination factors examined, high C-reactive protein (CRP) level was a significant predictor of unfavorable OS [hazard ratio (HR)=10.115, 95% confidence interval (CI)=2.447-41.806, P=0.001]. Among the post-vaccination factors, high CRP level and low frequency of CD3?CD26? cells were significant predictors of unfavorable OS (HR=23.127, 95% CI=2.919-183.233, P=0.003; HR=0.952, 95% CI=0.917-0.989, P=0.012). Taken together, our results suggest the feasibility of PPV for the treatment of refractory NSCLC. Evaluation of the identified factors before or at an early stage of vaccination could be potentially useful for selecting NSCLC patients who would likely have better prognosis following PPV.