Mouse embryonic diastema region is an ideal site for the development of ectopically transplanted tooth germ.

The anterior eye chamber and the kidney capsule of the mouse have been traditionally used for long-term culture of tooth germ grafts. However, although these sites provide an excellent growth environment, they do not represent real in situ sites for the development of a grafted tooth germ. Here, we describe a protocol to transplant a tooth germ into the mandibular diastema region of mouse embryos using exo utero surgery. Our results demonstrate that the mouse embryonic diastema region represents a normal physiological environment for the development of transplanted tooth germs. Transplanted tooth germs developed synchronically with and became indistinguishable from the endogenous ones. These ectopic teeth were vascularized and surrounded with nerve fibers, and were able to erupt normally. Thus, the exo utero transplantation approach will provide a new avenue to study tooth development and regeneration.     

Restricted expression of transgenic HLA-DRA gene in thymic epithelial cells and its role in acquisition of T cell tolerance to self-superantigens and processed DRα-derived peptide

We have established a set of transgenic mouse lines in which the HLA-DRA gene was expressed in different cell types. In one line (DRα-24), DRαEβ^b molecules were expressed on thymic medullary and cortical epithelial cells and all lineages of bone marrow-derived antigen-presenting cells (APC) except for thymic macrophages. By contrast, expression of the molecules in another line (DRα-30) was found on thymic medullary and cortical epithelial cells but not on bone marrow-derived APC in the thymus and periphery. To evaluate the role of thymic epithelial cells in acquisition of T cell tolerance, comparative analysis of DRα-24 and DRα-30 was performed. In DRα-30, T cells expressing TcR Vβ5 and Vβ11 were eliminated to comparable levels to those in DRα-24, suggesting that expression of the DRαEβ^b molecules on thymic epithelial cells are sufficient for clonal deletion of the self-superantigen-reactive T cells. In addition, CD4⁺ T cells from DRa-30 as well as those from DRα-24 were tolerant to DRα-derived peptide/I-A^b complex expressed on spleen cells from DRα-24 even in the presence of exogenous interleukin-2. These observations suggest that expression of the DRα chain in thymic epithelial cells could induce T cell tolerance directed toward naturally processed DRα-derived peptide bound to I-A^b molecules, probably via clonal deletion of the self-reactive T cells.     

Otx1 function overlaps with Otx2 in development of mouse forebrain and midbrain

Background: We previously reported that the homozygous mutation of Otx2 gene, a mouse cognate of the Drosophila head gap gene orthodenticle , causes failure in the development of the rostral head anterior to rhombomere 3, which may correspond to earlier Otx2 expression in cells destined for the anterior mesoendoderm. At the same time, the Otx2 heterozygous mutation displayed a phenotype characterized as otocephaly, probably related to expression in the anterior neuroectoderm at the subsequent pharyngula stage. Defects were characteristic in the most anterior and posterior regions of Otx2 expression where Otx1 , another mouse cognate of orthodenticle , is not or weakly expressed. They were not found in the region where Otx1 is expressed.
Results: In the present work, Otx1 null mutant mice were generated by gene targeting in embryonic stem cells. No defects were apparent in the regionalization of the early embryonic rostral brain. The newborn brain defects were subtle and most likely related to later Otx1 -unique expression. Otx1 and Otx2 double heterozygous mutant brains, however, exhibited marked defects throughout the fore- and midbrains, where defects were not apparent with a single mutation alone.
Conclusions: Otx1 and Otx2 play synergistic roles in the development of the forebrain and midbrain where both genes are expressed.     

急性胰腺炎发病机制和治疗的实验研究进展

在致病因素的作用下，各种消化酶和细胞因子、活性氧簇、内毒素、NF-κB等炎症介质的生成增加以及它们之间的相互作用，共同造成胰腺和远处器官的损伤。急性胰腺炎的治疗以一般治疗、抗氧化剂、生长抑素、糖皮质激素治疗为主，NBD多肽是研究的重点。     

Evaluation of the clinical pathway for laparoscopic cholecystectomy and simulation of short-term hospitalization.

The effectiveness of the clinical pathway for laparoscopic cholecystectomy was evaluated, and the efficiency of medical care was analyzed. The duration of hospitalization and the number of National Health Insurance (NHI) points for medical service fees were compared between 86 patients treated after introduction of the clinical pathway (pathway group) and 56 patients treated before introduction of the clinical pathway (pre-pathway group). In the pathway group, variance from the pathway occurred in 24 patients (27.9%) due to postponement of discharge in 7 patients, to earlier discharge in 5 patients, and to insertion of a bile duct catheter in 5 patients. Total and postoperative hospitalization times were significantly shorter in the pathway group than in the pre-pathway group (8.0 +/- 1.6 vs 13.7 +/- 9.0 days, p<0.0001, 5.4 +/- 1.1 vs 6.5 +/- 2.2 days, p<0.0001, respectively). In the pathway group, the total number of NHI points was lower and the number of points per day was higher. By simulation, the total number of NHI points for the 5-day pathway (discharge on postoperative day 3 or earlier) was significantly lower than that for the current 7-day pathway. Moreover, the weekly profit per bed with the 3-day pathway (discharge on postoperative day 1) was more than twice that with the current pathway. The results suggest that the clinical pathway for laparoscopic cholecystectomy is beneficial for patients and useful for the introduction of diagnosis procedure combination in our hospital.     

The dopamine agonist cabergoline provides neuroprotection by activation of the glutathione system and scavenging free radicals

Free radicals are involved in the pathogenesis and/or progression of Parkinson's disease (PD). Several ergot derivative dopamine (DA) agonists have been reported to scavenge free radicals in vitro and to show a neuroprotective effect in vivo. We investigated the in vitro free radical scavenging and antioxidant activities of cabergoline, a long-acting ergot DA agonist, as well as its ability to activate glutathione (GSH), catalase (Cat) and superoxide dismutase (SOD) activating effects and its in vivo neuroprotective properties against 6-hydroxydopamine (6-OHDA) intracerebroventricularly (i.c.v.) in mice. The striatal DA turnover induced by i.c.v. injection of 6-OHDA was completely normalized by pretreatment with cabergoline. Moreover, cabergoline scavenged free radicals in vitro and significantly reduced lipid peroxidation in vitro and in vivo. Furthermore, daily administration of cabergoline to mice significantly increased striatal GSH levels by activation of RNA expressions of GSH-related enzymes, although striatal Cat and SOD activities did not change. In addition, our present results suggest that repeated administration of cabergoline attenuates both 6-OHDA-induced nigrostriatal DAergic dysfunction and DA neuronal cell death, since cabergoline also had a neuroprotective effect in the immunohistochemical experiment. In conclusion, our findings indicate that the multiple antioxidant mechanisms of cabergoline, such as activation of the GSH system and the direct free radical scavenging activity, may explain the neuroprotective effect of this ergot DA agonist.     

Construction of Canine Herpesvirus Vector Expressing Foreign Genes Using a lacZ-TK Gene Cassette as a Double Selectional Marker

An improved method for constructing canine herpesvirus (CHV) recombinants expressing foreign genes by using the lacZ-TK gene cassette as a double selectional marker was developed. A recombinant CHV carrying the lacZ-TK gene at a targeted gene locus was constructed and used as a parental virus for generating new recombinants. The parental virus formed blue plaques and was sensitive to TK-specific drugs, while newly generated recombinants, in which the lacZ-TK gene was replaced with the desired foreign gene, become both resistant to the TK-specific drugs and formed white plaques. Recombinants were isolated by using the combination of drug selection and color selection. This improved method allows construction of recombinant CHV with great ease, because the drug selection can enrich the frequency of recombinant CHV from 0.01–0.1% to 10–80%. This method was employed to construct a recombinant CHV that expressed rabies virus (RV) glycoprotein (G protein). This revised version was published online in July 2006 with corrections to the Cover Date.     

Lethal toxicity of arthropod iridoviruses to an amphibian,Rana limnocharis

Summary Intraperitoneal injection of large doses of the two arthropod iridoviruses,Armadillidium iridescent virus (AvIV) andChilo iridescent virus, was lethal to the frog,Rana limnocharis. The LD₅₀ of purified AvIV for the frog was 2.7 µg. While UV-inactivated AvIV produced a lethal toxicity to the frog, heat-inactivated AvIV did not show any toxicity. Lethal toxicity of AvIV was neutralized by the homologous rabbit antiserum but not by the heterologous antisera.     

Ex vivo expansion and prophylactic infusion of CMV-pp65 peptide-specific cytotoxic T-lymphocytes following allogeneic hematopoietic stem cell transplantation.

Cytomegalovirus reactivation and infection post-allogeneic hematopoietic stem cell transplant continue to cause morbidity and mortality. Current pharmacologic therapies are limited by side effects. Adoptive transfer of ex vivo generated cytomegalovirus-specific T cells has the potential to restore immunity, prevent cytomegalovirus, and circumvent the need for pharmacologic therapies. We have generated donor-derived cytomegalovirus-specific cytotoxic T cells using dendritic cells pulsed with the HLA-A2 restricted nonapeptide NLVPMVATV (NLV) derived from the cytomegalovirus-pp65 protein. These cytotoxic T cells have been given prophylactically to 9 recipients aged 4 to 65 years on or after day 28 post-allogeneic hematopoietic stem cell transplant. Only 2 of 9 recipients received T cell depletion in vivo or in vitro. There were no immediate adverse reactions to the infusions. During 97-798 days of follow-up, 2 recipients developed cytomegalovirus reactivation; neither developed cytomegalovirus disease or required pharmacotherapy. Three recipients developed acute graft versus host disease after infusion. Two recipients died, 1 from thrombotic thrombocytopenia purpura secondary to cyclosporine, 1 from complications of graft versus host disease. A transient increase in numbers of cytomegalovirus-specific T cells demonstrated by NLV-tetramer binding was seen in 6 recipients. Prophylactic adoptive transfer of NLV-specific T cells is safe and may be effective in preventing cytomegalovirus reactivation.