Selection of stereotyped VH81X-{micro}H chains via pre-B cell receptor early in ontogeny and their conservation in adults by marginal zone B cells

The pre-B cell receptor (preBCR) plays critical roles in early B cell differentiation. It has been shown that not all muH chains are capable of pairing with surrogate light (SL) chains to form preBCR. Here, we established a novel system to differentially identify two types of early pre-B cell populations in bone marrow and fetal liver of mice, one producing SL-pairing muH chains and the other producing SL-non-pairing muH chains. The former population accounted for 80% of all the early pre-B cells in adult bone marrow, while it accounted for only 20% of those in fetal liver. Comparison of the two types of pre-B cell populations in fetal liver revealed the structural difference between SL-pairing and -non-pairing muH chains encoded by the V(H)81X segment that was most frequently utilized in fetal liver pre-B cells but rarely expressed by B cells generated in adults. PreBCR played an important role in the positive selection of V(H)81X-muH chains carrying the characteristic sequences of the complementarity-determining region 3 with little or no nibbling or N nucleotide addition, leading to their predominance in neonatal splenic B cells. These fetal-type V(H)81X-muH chains were also detected in adult spleen, but almost exclusively in marginal zone (MZ) B cells in contrast to the adult-type V(H)81X-muH chains. This strongly suggests that neonatally generated and selected B cells expressing the stereotyped V(H)81X-muH chains are maintained in the adult MZ and could function as innate-like lymphocytes.     

Mutational screening of APP gene in patients with early-onset Alzheimer disease utilizing mismatched PCR-RFLP

To elucidate the frequency of mutations of the β/A4 amyloid protein precursor (APP) gene in early-onset Alzheimer disease, we designed a mismatched PCR-RFLP that can identify all kinds of missense mutations at codon 717 in addition to the seven kinds of known mutations at exon 17. When we screened mutations at exon 17 utilizing this method and the double missense mutations at exon 16 of the APP gene by PCR-RFLP, no cases revealed mutations of the APP gene among 13 familial and 54 sporadic cases, except one family (OS-1) that had previously been reported and used as a positive control of APP717(Val → Ile). Our results support the hypothesis that mutations in the APP gene are not major causes in early-onset Alzheimer disease.     

Rapid diagnosis of human herpesvirus 6 infection by a novel DNA amplification method, loop-mediated isothermal amplification

A novel nucleic acid amplification method, termed loop-mediated isothermal amplification (LAMP), which amplifies DNA with high specificity, efficiency, and rapidity under isothermal conditions, may be a valuable tool for the rapid detection of infectious agents. LAMP was developed for human herpesvirus 6 (HHV-6), and its reliability was evaluated in this study. Although LAMP products were detected in HHV-6 B and HHV-6 A DNA, they were not detected in HHV-7 and human cytomegalovirus DNA. The sensitivity of the original HHV-6 LAMP protocol was 50 copies/tube. In order to increase the method's sensitivity, HHV-6 LAMP was modified by increasing the primer concentration. As a result of the modification, sensitivity increased to 25 copies/tube. After these initial validation studies, 13 patients with fever were tested for HHV-6 by viral isolation, serological analysis, and HHV-6 LAMP. In three of the eight patients with primary HHV-6 infection, HHV-6 DNA was detected in whole blood by the original HHV-6 LAMP protocol in not only the acute phase but also the convalescent phase. HHV-6 DNA was detected by modified HHV-6 LAMP in all eight plasma samples collected in the acute phase; however, no HHV-6 DNA was detected in plasma samples collected in the convalescent phase. Although HHV-6 DNA was detected in both the acute and convalescent phases of whole-blood samples in patients with past HHV-6 infection, it was not detected in plasma samples that did not contain latent viral DNA. Thus, detection of HHV-6 DNA in plasma by using this modified HHV-6 LAMP protocol is appropriate for diagnosis of active HHV-6 infection.     

Dominant-Negative Rho, Rac, and Cdc42 Facilitate the Invasion Process of Vibrio parahaemolyticus into Caco-2 Cells

To clarify the invasive process of Vibrio parahaemolyticus, an invasion assay was performed using cells expressing dominant negative small GTPases of the Rho family. This assay showed that the dominant negative host phenotype facilitates bacterial invasion, suggesting that the mechanism of V. parahaemolyticus invasion differs from that reported for other invasive bacteria.     

Histopathological studies on virulence of dipeptidyl aminopeptidase IV (DPPIV) of Porphyromonas gingivalis in a mouse abscess model: use of a DPPIV-deficient mutant

To elucidate the role of dipeptidyl aminopeptidase IV (DPPIV) in the virulence of Porphyromonas gingivalis, mice were infected with either a wild-type strain or a DPPIV-deficient mutant using an abscess model. Histopathological analysis of the resulting lesions indicated that DPPIV participates in virulence through the destruction of connective tissue and the less effective mobilization of inflammatory cells.     

Pathogenesis of Rinderpest Virus Infection in Rabbits I. Clinical Signs, Immune Response, Histological Changes, and Virus Growth Patterns

Rabbits were intravenously inoculated with an attenuated rinderpest virus (L strain), and general patterns of the disease were investigated. The rabbits developed fever with concomitant occurrence of diarrhea and lymphopenia. Early production of interferon was followed by a rise of neutralizing antibody. Histological examinations revealed an involvement of all of the lymphoid tissues, with primary lesions consisting of necrosis of the lymphoid follicles and formation of giant cells. Immunofluorescent examinations suggested that the virus growth was present in almost all of the lymphoid tissues. The possibility of application of this experimental system for the study of systemic infection by measles virus was discussed.     

Renal primitive neuroectodermal tumor: An immunohistochemical and cytogenetic analysis

The cytogenetic and morphologic characteristics of a case with a primitive neuroectodermal tumor (PNET) arising from the left kidney in a 22 year old man are presented. The patient was detected as having a left renal mass with a tumor embolus In the inferior vena cava and multiple pulmonary metastases. A radical nephrectomy with tumor embolectomy of the Inferior vena cava, along with a resection of the pulmonary nodules were performed. Histologic examination revealed a dense proliferation of small round cells with many Homer-Wright type rosettes and perlvascular pseudo-rosettes. Immunohlstochemically, the tumor cells stained strongly positive for HBA71(p30/32^{M IC2}), a surface glycopro-teln specific to PNET and Ewlng's sarcoma. In addition, the tumor cells expressed several neural markers (neuron specific enolase, neurofilament, synaptophysin, and Leu-7) and vimentin, while the epithelial, muscular, and lymphocytlc markers were negative in the tumor cells. Cytogenetic analysis of cultured tumor cells showed a reciprocal translocation t(11; 22)(q24; q12) that is considered to be specific to PNET and Ewing's sarcoma. In conclusion, this case suggested that a karyotyping analysis is a useful diagnostic tool for renal PNET, and it may therefore be utilized to help distinguish between difficult cases of small round cell tumors and Wilms' tumor of the kidney.