Extracellular Vesicles of Stem Cells to Prevent BRONJ.

Extracellular vesicles (EVs), several tens to hundreds of nanometers in size, are vesicles secreted by cells for intercellular communication. EVs released from mesenchymal stem cells (MSC-EVs) have the potential to treat multiple diseases. This study aimed to determine the effects of MSC-EVs on bisphosphonate-related osteonecrosis of the jaw (BRONJ), whose pathogenesis and treatment are not yet established. To this end, zoledronic acid (ZOL) was administered to bone marrow cells and fibroblasts in vitro. In vivo, a BRONJ model was produced by administering ZOL to rats and extracting teeth. Each MSC-EV-treated and nontreated group was compared histologically and molecularly. In vitro, the nontreated group showed an increased number of ß-galactosidase-positive cells and expression of senescence-associated genes p21, pRB and senescence-related inflammatory cytokines. Conversely, MSC-EV administration decreased the number of senescent cells and expression levels of p21, pRB and inflammatory cytokines. In vivo, in the nontreated group, the socket was partially uncovered by the oral epithelium, leaving an exposed bone. Conversely, in the MSC-EV-treated group, the socket was healed. Besides, in the nontreated group, ß-galactosidase-positive cells existed in the socket and colocalized with the CD90 and periostin-positive cells. However, there were few ß-galactosidase-positive cells in the MSC-EV-treated group. Furthermore, gene expression of stem cell markers Bmi1 and Hmga2 and the vascular endothelial marker VEGF was significantly increased in the MSC-EV-treated group, compared with that in the nontreated group. These results indicate that MSC-EVs prevent ZOL-induced senescence in stem cells, osteoblasts, and fibroblasts and reduce inflammatory cytokines. Furthermore, administration of MSC-EVs prevented senescence of cells involved in wound healing and the spread of chronic inflammation around senescent cells, thereby promoting angiogenesis and bone regeneration and preventing BRONJ.

A technique for complete thymectomy in adult rats.

50 open thymectomies were performed in adult rodents using intubation combined with a fibrin glue able to prevent hemorrhage and pulmonary air leakage. This method had a 100% success rate and lower mortality than the ordinal suction procedure. Although the conventional suction thymectomy has been widely used, the open thymectomy method would permit more complete thymectomies for immunological studies.

Prevention by liver transplantation of the graft-versus-host reaction and allograft rejection in a rat model of small bowel transplantation.

In the rat combination of DA (MHC haplotype RT1av1) donor into PVG(RT1c) recipient, liver grafts are not rejected and allow the acceptance of other organ grafts from the same donor strain. Here we show that an existing DA liver graft allows the acceptance of a DA small bowel graft in the PVG; the liver graft also prevented the graft-versus-host reaction normally associated with small bowel grafting. In addition, a liver graft could suppress the GVHR in F1 hybrid recipients of parental lymphoid cells, a classic GVHR model. GVHR suppression was immunologically specific and required that donor lymphocytes and liver be of the same strain. Besides their clinical implications, these results demonstrate the capacity of the liver for tolerance induction and suggest that it may play a physiological role in negative selection of T cells.

Evidence that the elevation of soluble MHC class I antigens in the serum precedes the onset of graft-versus-host disease and is correlated with the severity of the disease in rats.

We examined the changes in the levels of soluble major histocompatibility antigen complex (MHC) class I antigens in the serum under a lethal or nonlethal state of graft-versus-host-disease (GVHD) induced by injecting various doses of PVG rat splenic lymphocytes into (DA x PVG)F1 rats. All rats receiving 4 x 10(8) lymphocytes (lethal dose) died on day 20-36 showing typical features of GVHD, while the injection of 4 x 10(7) cells (nonlethal dose) induced no sign of GVHD. When rats were inoculated with a nonlethal dose of lymphocytes prior to the injection of a lethal dose, all rats survived with or without showing transient GVHD. Preceding the onset of GVHD the levels of soluble class I antigens increased significantly to 1094 +/- 487 ng/ml (mean +/- SD, n = 4) from 3 days after the injection of a lethal dose to the time of death, whilst the levels in the nonlethal dose group remained unchanged. Rats with transient GVHD in the preinoculated group showed the increase of soluble class I antigens to the same extent as rats with lethal GVHD, suggesting that GVHD was systemically ongoing. The levels of soluble class I antigens also correlated with the severity of GVHD as judged by daily observation and histological studies. Rats receiving a lethal dose showed destructive alteration of spleen structure and cellular infiltration in the portal area of the liver before the animals started to show signs of GVHD, whereas rats in the nonlethal dose group exhibited no marked change. These data suggest the possibility of serum soluble class I antigens being not only a diagnostic but also a prognostic marker for GVHD.

Fulminant second-set allograft rejection and endoscopic findings following small bowel transplantation in the rat.

In the presensitized recipient who has been exposed to donor antigens, second-set rejection takes the form of severe hyperacute graft rejection. Second-set allograft rejection was studied following small bowel transplantation in the rat. Heterotopic intestinal grafting was performed from DA (RT1a) donors to PVG (RT1c) recipients 4 weeks after DA skin sensitization. The endoscopic images and histological specimens were compared with those of syngeneic and first-set rejected grafts. Endoscopically, diffuse erosions of the graft were detected from day 1. Mucosal necrosis progressed rapidly, and was accompanied by massive bleeding on days 3-5. These findings were similar to the course of severe necrotizing hemorrhagic enteritis. Histologically, interstitial edema and hemorrhage with massive infiltrations of neutrophils were manifested from day 1. Mesenteric vessels were completely occluded by thrombi on days 3-5. The grafted intestine had became totally necrotic by day 5. Microscopic findings strongly suggested that destructive graft necrosis was due to vascular damage caused by humoral factors. All the presensitized rats (n = 11) died showing systemic septic signs by day 11 after small bowel transplantation. We conclude that lethal hyperacute rejection occurred in presensitized recipients, even when the graft was transplanted heterotopically. Endoscopic evaluation is beneficial for the early diagnosis of graft rejection. Immediate graft removal should be mandatory as a rescue treatment in second-set rejection of the small intestine.

Endoscopic findings in transplanted allo-intestine of rats after discontinuance of immunosuppressive agent.

In clinical practice, graft rejection in small-bowel transplantation should be diagnosed before irreversible condition of the graft. We have already reported the usefulness of endoscopic examination for the early detection of acute rejection in a rat model. Here we evaluated rejection after discontinuance of methyl-deoxyspergualin by endoscopy. Heterotopic small-bowel transplantation was performed by the cuff method from a DA to a LEW rat. Endoscopic and histological examinations were performed through the stomas. Two-week administration of methyl-deoxyspergualin significantly prolonged graft survival. Graft rejection after discontinuance of the agent occurred much more slowly than rejection without the immunosuppressive drug. Erosive mucosal changes were endoscopically observed in the early phase of rejection in rats that did not receive the immunosuppressant. However, endoscopic findings after discontinuance of methyl-deoxyspergualin indicated edematous changes and thickening of the wall without erosion, and, histologically, the grafted intestine showed slowly-progressing rejection with flattened villi. If we pay attention to edematous changes and hardening of intestinal wall, and take selective biopsies, endoscopic examination may improve the early diagnosis of slowly progressive rejection in the clinical setting.

Segmental small bowel transplantation in the rat: comparison of lipid absorption between jejunal and ileal grafts.

BACKGROUND: From the immunological point of view, it is very important to evaluate the efficacy of segmental small bowel transplantation and to determine which part of the intestine, jejunum, or ileum should be used. In the present study, we investigated the absorptive function of the transplanted jejunum and ileum in the rat. METHODS: Syngeneic segmental small bowel transplantation (jejunal or ileal grafting) was performed. After surgery, body weight and fecal fat excretions were measured. In addition, bile acid concentration in bile juice was measured, and the response of both serum lipase and bile flow rate after oleic acid stimulation was evaluated. Recanalization of the lymphatic vessels was investigated by lymphangiography. RESULTS: There was no significant difference in body weight change between normal controls, jejunum-transplanted rats, and ileum-transplanted rats. In short gut rats, however, body weight was significantly impaired. Fecal fat excretion in short gut rats was the highest in the four groups, and it was significantly lower in ileal grafts than in jejunal grafts. Ileum-transplanted rats also showed a significantly higher bile acid concentration in bile juice than jejunum-transplanted rats. After oleic acid stimulation, serum lipase responded better in ileal transplants than in jejunal transplants, but the bile flow rate did not show significant change in either group. Recanalization of the lymphatic vessels was established on the 28th postoperative day. CONCLUSIONS: These results clearly show that ileal transplantation is more conducive to lipid absorption than jejunal transplantation.

Natural history of a patient with malignant nonfunctioning islet cell tumor associated with unresectable multiple liver metastases.

Islet cell tumors of the pancreas are uncommon. Approximately 15% of islet cell tumors are nonfunctioning and have a higher malignancy rate than their functioning counterparts. Though, because of the rarity of malignant nonfunctioning islet cell tumors, the natural history of a patient with this tumor has not been clearly defined. We describe a young patient with unresectable malignant nonfunctioning islet cell tumor associated with multiple liver metastases. He was treated with palliative therapies to improve his quality of life, but did not undergo surgical removal of tumors or systemic chemotherapy. He survived for 46 months since laparotomy for histological diagnosis. Our findings may represent the natural history of patients with unresectable malignant nonfunctioning islet cell tumor, and suggest that palliative therapy may contribute not only to the improvement of a patients' quality of life but also the prolongation of survival.

Small bowel transplantation for pediatric short bowel syndrome: evaluation of the graft length required for development and the immunologic aspects relating to graft length.

Small bowel transplantation (SBT) is thought to be the only radical treatment for short bowel syndrome in childhood. It is very important that the length of the graft and the type of intestine be chosen carefully because this will determine the outcome of transplantation. This model of short bowel syndrome in the rat confirms that total intestinal resection results in malnutrition and failure to gain weight. After 10 cm of ileum was transplanted orthotopically in a syngeneic combination in rats with total intestinal resection, the animals gained weight. The authors determined that 10 cm of terminal ileum is the minimum length required for survival. Second, the immunologic basis of lethal graft-versus-host reaction (GVHR) as it relates to the intestinal graft length was also evaluated. Ileal grafts of 10 cm from LEW (RT1l) rats were implanted heterotopically into (LEW x BN)F1 rats. Ileal grafts of 10 or 40 cm were implanted from BN(RT1n) rats into (LEW x BN)F1 animals. A lethal GVHR always occurred after grafting a 10 cm ileum in the LEW/F1 combination, whereas only 17% of the BN/F1 recipients died of typical GVHR. However, in the latter combination, 67% lethal GVHR was induced when 40 cm of the intestinal graft was implanted. These results indicate that mesenteric lymph nodes are a major source of lethal GVHR, but gut-associated lymphoid tissue can also induce this.

The effect of additional autopolyploidization in a slow growing cellulase hyperproducer of Trichoderma.

M14-2 is a cellulase hyperproducer derived from Trichderma reesei QM 6a, but with a growth rate lower than that of the original strain. When M14-2 was autopolyploidized followed by haploidization and selection, the strain with both a higher cellulase productivity per mycelia and a higher growth rate could be obtained as M14-2B. This strain seemed to be constructed using gene sources amplified by additional autopolyploidization.

Successive construction of cellulase hyperproducers of Trichoderma using hyperpolyploids.

When the swollen conidia of Trichoderma reesei QM 6a are treated with 0.1% (w/v) colchicine solution, huge autopolyploid nuclei can be formed in those swollen conidia. When a mycelial mat derived from such a conidum is treated with a haploidizing reagent, benomyl, many fan-shaped sectors are produced from the colony, and cellulase hyperproducers are selected from conidia on the colony. When colchicine and benomyl treatments are repeated on cellulase hyperproducers, new hyperproducers can be constructed successively and systematically. Moreover, when conidia derived from autopolyploids are treated with ethylmethanesulfonate solution, another type of cellulase hyperproducers (polyploids) can be obtained.

[A pharmacokinetic study of cefoperazone during percutaneous transhepatic cholangial catheterization].

The metabolic fate of cefoperazone (CPZ) was studied in 19 cases which underwent percutaneous transhepatic cholangial catheterization (PTC-catheterization, PTCC) and were under various conditions of the liver function. The peak of bile levels of CPZ immediately after PTCC differed greatly from one case to another at 12.6-7,260 micrograms/ml with 1 g intravenous injection and 23.0-5,800 micrograms/ml with 2 g intravenous injection. The ratio of the peak of bile level to the serum level immediately after PTCC showed the highest negative correlation with the serum total bilirubin level. It also showed a significant negative correlation with GOT, GPT, Al-P and LAP. The serum CPZ level and half-life showed no significant trend except half-life showed a significant correlation with LAP. The recovery rate in urine up to 12 hours was in the range of 14.8-93.6%, showing a significant correlation with the ratio of the peak of bile levels to the serum level and the date of liver function tests. The bile level, serum level and recovery rate in urine at the time the bile outflow from the catheter has become constant after PTCC (during the course of PTCC) showed a trend almost similar to that immediately after PTCC, there being no significant difference as to each parameter during the course of PTCC and immediately after PTCC. In the cases in which the sample was collected by the cross-over technique, the ratio of the peak of bile levels to the serum level from immediately after PTCC to during the course of PTCC increased in 2 cases and decreased in 6 cases. The 2 cases that showed the increase in the ratio were the case in which the serum total bilirubin level improved almost to normal. Findings above suggest that sufficient biliary decompression can improve the movement of CPZ into bile, despite the fact that the pharmacokinetics of CPZ is affected by the liver function, particularly serum total bilirubin level, that a decrease in the movement to bile and a compensatory increase in urinary excretion are observed in jaundice and disturbance of the liver function and that the ratio of the peak of bile level to the serum level decreases during the course of PTCC rather than immediately after PTCC in some cases.

[Awareness of health and welfare planning for elderly and methods to increase awareness: based on social marketing].

To clarify the socio-psychological factors affecting awareness of health and welfare planning for the elderly, a survey of community residents was performed in the Tokyo metropolitan area. The results obtained were as follows: 1. There were three factors which affected awareness concern about health and welfare services for the elderly as a social issue: concern about the local community; and concern about the local government. 2. Two other factors which did not affect the level of awareness were: anxiety about the health and welfare services for the elderly as a personal issue; and having personal experience of nursing care. From these results, a method to increase awareness was studied based on social marketing methods. Two primary target groups for increasing awareness were identified. One target was people who have the socio-psychological factors described above. Another target was people who have the potential need for health and welfare services but who were not aware of it. That is, they have relatively high anxiety about health and welfare services for the elderly as a personal issue and have personally experienced nursing care. The method of approach for these targets were studied. For the first group, the amount of the information available seemed to be important because these persons are ready to recognize the need for planning. Therefore, efficient information channels should be selected. For the second group, approaches that generate greater consumer participation by presenting this as an efficient method for solution of the problem should be adopted.

[Mental development of 'meanings' related to scripts: 'dining' script as an example].

The purpose here is to investigate how the process of relating 'meanings' to scripts is influenced by the developmental changes of concepts about these activities, with an example of 'dining'. Based on previous studies that the concept of 'dining' develops from physiological function orientation to social function orientation, second, fourth, and sixth graders, and college students were asked to plan 'dining' with a physiological goal or a social one. The results are as follows. (1) Generated plans became more connected with goals developmentally in both social and physiological context. (2) Second graders who attached much importance to physiological function of 'dining' were apt to generate plans with physiological meanings even in social goal context.