[Causes and background of death in elderly patients with advanced dementia].

AIM: To examine the causes of death in elderly patients with advanced dementia, we retrospectively investigated the medical records for death discharge cases hospitalized in the Department of Geriatric Medicine at Saiseikai-Nakatsu Hospital and examined death certificates issued throughout the hospital. METHODS: (1) From 2010 to 2013, 31 patients with advanced dementia died in the hospital or were discharged to receive terminal care at home. We evaluated their medical records to examine the pathological background and disease with which they were diagnosed when admitted to and discharged from the hospital. (2) In order to assess the relationship between disease and dementia, we examined death certificates with "senility" or " (aspiration) pneumonia" recorded as the direct death cause issued throughout the hospital in the one-year period of 2013. RESULTS: (1) There were many cases in which eating problems and dysphagia influenced the clinical course. A total of 21 patients died from eating problems and/or dysphagia. (2) All 13 cases with "senility" recorded as the direct death cause on the death certificate involved severe dysphagia. Investigating the medical records, 11 patients had advanced dementia and two patients had end-stage Parkinson's disease. In total, 46 cases were diagnosed as involving " (aspiration) pneumonia", whereas there were no cases in which the records mentioned dementia or dysphagia in another column on the death certification. CONCLUSIONS: Advanced dementia is a mortal illness, and most patients with advanced dementia have dysphagia. Clinicians should be aware of the fact that dysphagia may lead to aspiration pneumonia and is a significant cause of death. Understanding the clinical course of dementia is important for determining the cause of death.

Bleeding from the small intestine and aortic regurgitation in Noonan syndrome.

A 62-year-old man was admitted to our hospital because of melena. On admission physical examination revealed that he had typical features of Noonan syndrome (NS). Investigation via upper endoscopy with the single balloon demonstrated oozing from the small intestine. Bleeding sometimes occurs in patients with NS. We speculated that coagulation defects or vascular malformations might have been present at the first visit in this case. However, coagulation function was normal. By upper endoscopy with the single balloon we clearly revealed the angioectasia in the small intestine. This case documents the first association among NS, aortic regurgitation and angioectasia in the small intestine.

Administration of a determinant of preproinsulin can induce regulatory T cells and suppress anti-islet autoimmunity in NOD mice.

Antigen-specific immunotherapy is expected to be an ideal strategy for treating type 1 diabetes (T1D). We investigated the therapeutic efficacy of a peptide in the leader sequence of preproinsulin, which was selected because of its binding affinity to the MHC I-A(g7) molecule. Preproinsulin-1 L7-24 peptide (L7-24) emulsified in Freund's incomplete adjuvant was administered subcutaneously to NOD mice. Administration of L7-24 increased the proportion of regulatory T cells in the spleen. Splenocytes of NOD mice immunized with this peptide secreted IL-4 and IL-10 in response to L7-24. This peptide also significantly prevented the development of diabetes and cured some newly diabetic NOD mice without recurrence. L7-24 peptide, which has a high affinity for pockets of I-A(g7), induced regulatory T cells and showed therapeutic effects. This peptide may provide a new approach for developing antigen-specific immunotherapy for autoimmune diabetes.

Regulatory CD8+ T cells induced by exposure to all-trans retinoic acid and TGF-beta suppress autoimmune diabetes.

Antigen-specific regulatory CD4(+) T cells have been described but there are few reports on regulatory CD8(+) T cells. We generated islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP)-specific regulatory CD8(+) T cells from 8.3-NOD transgenic mice. CD8(+) T cells from 8.3-NOD splenocytes were cultured with IGRP, splenic dendritic cells (SpDCs), TGF-beta, and all-trans retinoic acid (ATRA) for 5days. CD8(+) T cells cultured with either IGRP alone or IGRP and SpDCs in the absence of TGF-beta and ATRA had low Foxp3(+) expression (1.7+/-0.9% and 3.2+/-4.5%, respectively). In contrast, CD8(+) T cells induced by exposure to IGRP, SpDCs, TGF-beta, and ATRA showed the highest expression of Foxp3(+) in IGRP-reactive CD8(+) T cells (36.1+/-10.6%), which was approximately 40-fold increase compared with that before induction culture. CD25 expression on CD8(+) T cells cultured with IGRP, SpDCs, TGF-beta, and ATRA was only 7.42%, whereas CD103 expression was greater than 90%. These CD8(+) T cells suppressed the proliferation of diabetogenic CD8(+) T cells from 8.3-NOD splenocytes in vitro and completely prevented diabetes onset in NOD-scid mice in cotransfer experiments with diabetogenic splenocytes from NOD mice in vivo. Here we show that exposure to ATRA and TGF-beta induces CD8(+)Foxp3(+) T cells ex vivo, which suppress diabetogenic T cells in vitro and in vivo.

[Anti-aging medicine: the evidence to the value of the antihypertensive drugs, hypoglycemic drugs and statins].

Aging is associated with a large increase in the prevalence and incidence of arteriosclerotic diseases including cerebrovascular disease and coronary artery disease. Prevention of arteriosclerosis is a major challenge in order to increase longevity of populations. Hypertension, diabetes and dyslipidemia are known as lifestyle-related diseases and risk factors for arteriosclerosis. The tight control of blood pressure, glucose and LDL cholesterol is important in preventing arteriosclerosis. Many clinical trials have been revealed pleiotropic effects among antihypertensive drugs, hypoglycemic drugs and statins, and these effects are useful for anti-aging. Selection of appropriate medicines to manage the risk factors would be a way to prevent senescence.

NO-mediated cytotoxicity contributes to multiple low-dose streptozotocin-induced diabetes but not to NOD diabetes.

Type 1 diabetes (T1D) is caused mostly by autoimmune destruction of pancreatic beta-cells, the precise mechanism of which remains unclear. Two major effector mechanisms have been proposed: direct cell-mediated and indirect cytokine-mediated cytotoxicity. Cytokine-mediated beta-cell destruction is presumed mainly caused by NO production. To evaluate the role of iNOS expression in T1D, this study used a novel iNOS inhibitor ONO-1714. ONO-1714 significantly reduced cytokine-mediated cytotoxicity and NO production in both MIN6N9a cells and C57BL/6 islets in the presence of IL-1beta, TNF-alpha, and IFN-gamma. To evaluate whether NO contributes to diabetes progression in vivo, ONO-1714 was administered to four different mouse models of autoimmune diabetes: multiple low-dose STZ (MLDS)-induced C57BL/6, CY-induced, adoptive transfer and spontaneous NOD diabetes. Exposure to STZ in vitro induced NO production in MIN6N9a cells and C57BL/6 islets, and in vivo injection of ONO-1714 to MLDS-treated mice significantly reduced hyperglycemia and interestingly, led to complete suppression of cellular infiltration of pancreatic islets. In contrast, when ONO-1714 was injected into spontaneous NOD mice and CY-induced and adoptive transfer models of NOD diabetes, overt diabetes could not be inhibited in these models. These findings suggest that NO-mediated cytotoxicity significantly contributes to MLDS-induced diabetes but not to NOD diabetes.

One amino acid difference is critical for suppression of the development of experimental autoimmune diabetes (EAD) with intravenous injection of insulinB:9-23 peptide.

InsulinB:9-23 peptide (insB:9-23) reactive T cells has been reported as crucial for type 1 diabetes. In this study, experimental autoimmune diabetes (EAD) mice, which subcutaneous immunization of ins1 or 2B:9-23 induced autoimmune diabetes in F1(B7.1B6 x BALB/c), was investigated for antigen specific therapy to delete pathogenic T cells. Intravenous injection of ins1 or 2B:9-23 significantly delayed the development of diabetes on the corresponding peptide-induced EAD (ins1EAD or ins2EAD) concomitant with reduced insulitis and insulin autoantibodies expression. Population of Foxp3(+) CD4(+) T cell was unchanged whereas the level of anti-insB:9-23 specific IgG(2a) but not IgG(1) were specifically decreased, suggesting reduction of pathogenic insB:9-23 reactive T cells. Most interestingly, intravenous administration of ins2B:9-23, whose amino acid sequence had one amino acid difference at position 9 delayed the development of diabetes in both ins1EAD and ins2EAD whereas ins1B:9-23 administration delayed diabetes in the ins1EAD but not ins2EAD, suggesting that one amino acid difference gives critical influence on the effect of intravenous injection of antigenic peptide for type 1 diabetes.

Prevention of recurrent but not spontaneous autoimmune diabetes by transplanted NOD islets adenovirally transduced with immunomodulating molecules.

Pancreatic islet transplantation has the potential to maintain normoglycemia in patients with established type 1 diabetes, thereby obviating the need for frequent insulin injections. Our previous study showed that recombinant IL-12p40-producing islets prevented the recurrence of NOD diabetes. First, to see which immunomodulating molecule-secreting islet grafts can most powerfully prevent diabetes development in NOD mice without immunosuppressant, NOD islets were transfected with one of the following adenoviral vectors: Ad.IL-12p40, Ad.TGF-beta, Ad.CTLA4-Ig, or Ad.TNF-alpha after which they were transplanted under the renal capsule of acutely diabetic NOD mice. The immunomodulating molecules produced by these adenovirus-transfected islets in vitro were 74+/-19ng, 50+/-4ng, 821+/-31ng, and 77+/-18ng/100 islets, respectively. Transplantation of IL-12p40, TNF-alpha, and CTLA4-Ig but not TGF-beta-secreting islets displayed enhanced survival and delayed diabetes recurrence in recent-onset diabetic recipients. IL-12p40-producing islet grafts most powerfully prevented recurrent diabetes in NOD mice. In addition, local production of TNF-alpha and CTLA4-Ig significantly prolonged islet graft survival. In second series of experiment, these manipulated islets were transplanted under the renal capsule of 10-week-old NOD recipients and were also transplanted subcutaneously into 2-week-old NOD recipients. Transplantation of these islets into 2- or 10-week-old pre-diabetic mice failed to protect them from developing diabetes; in fact, transplantation of Ad.TNF-alpha-transfected islets into 2-week-old mice actually accelerated diabetes onset. Taken together, this approach was ineffectual as a prophylactic protocol. In conclusion, this study showed comparisons of the immunomodulating effects of 4 different adenoviral vectors in the same transplantation model and local production of IL-12p40, TNF-alpha and CTLA4-Ig significantly prevented recurrent diabetes in NOD mice.

Intravenous administration of proinsulin 1 or 2-expressing fiber-mutant recombinant adenovirus vector protects against the development of diabetes in NOD mice.

Insulin has been reported as a major autoantigen in both human and murine type 1 diabetes (T1D). Insulin1-knockout NOD mice with only insulin2 are protected against the development of autoimmune diabetes, suggesting that insulin1 has strong immunogenicity and insulin2 has weak immunogenicity or a possible protective role in the pathogenesis of type 1 diabetes. In this study, we have developed fiber-mutant adenovirus vectors that express murine proinsulin1 or proinsulin2 (named Ad.Pins1-RGD/Ad.Pins2-RGD) and administered those virus vectors to the NOD mouse to evaluate modulation of autoimmune responses. The intravenous administration of either Ad.Pins1-RGD or Ad.Pins2-RGD at 3 and 5 weeks of age strongly suppressed the development of overt diabetes, accompanied by a significant reduction of insulin autoantibody (IAA), and suppression of disease was similar between administration of Ad.Pins1-RGD and that of Ad.Pins2-RGD. Our study suggests that systemic administration of fiber-mutant adenovirus vectors, which induce transient expression of proinsulin, may be applicable to a gene therapy inducing tolerance to insulin.

Autoreactive T cell response in CD25-negative fraction of peripheral blood mononuclear cells in established type 1 diabetes.

CD4(+)CD25(+) T cells (Tregs) play a critical role in maintaining dominant peripheral tolerance, and pathogenic autoreactive T cells may be frequent in the CD25-negative fraction of peripheral blood mononuclear cells (PBMCs) from patients with autoimmune disease. We therefore investigated whether T cell autoimmune responses to recombinant GAD65 can be detected by the use of ELISPOT assay in the CD25-negative fraction of PMBCs from Japanese type 1 diabetes (T1D) patients. The frequency of CD4(+)CD25(+) T cells was not different among patients with newly developed T1D, established T1D, and healthy controls. The CD25 positive cell-depleted fraction was obtained by negative selection with antihuman CD25 magnetic beads, reducing the number of CD4(+)CD25(+) T cells from 4-5% to less than 1%. In whole PBMC fraction, there was a significant elevation of IFN-gamma spots in PBMCs from recently diagnosed patients with T1D (P < 0.05), whereas the number of IFN-gamma spots from patients with established T1D was not significant. In the CD25-negative fraction, unlike whole PBMCs, we observed the significant IFN-gamma spots to GAD65 in the fraction from patients with established T1D (P < 0.05), but not in those with recently diagnosed disease. The phenomena were not observed for IL-4 spots. Our data suggest a possible role of Tregs maintaining dominant peripheral tolerance in T1D and application of further improved T cell assay detecting autoimmunity even in established T1D.

Insulin as a T cell antigen in type 1 diabetes supported by the evidence from the insulin knockout NOD mice.

Rodents have two functional preproinsulin genes named insulin 1 and insulin 2 on different chromosome and have two amino acid differences in insulin B chain. We have established insulin 1 or insulin 2 knockout (KO) non-obese diabetic (NOD) colonies in the animal institute of Kobe University and evaluated anti-insulin autoimmunity. Similar to the previous report, insulin 1-KO provides strong protection from insulitis (islet-infiltration of mononuclear cells) and diabetes, whereas the insulin 2-KO markedly accelerated insulitis and development of diabetes even at further backcross breeding with NOD/Shi/Kbe mice (P<0.0001). Expression of serum anti-insulin autoantibodies (IAA) was enhanced in insulin 2-KO mice at a time between 10 and 15 weeks of age (P<0.005) while the expression of insulin 1-KO NOD mice was rather reduced. Furthermore, T cell reactivity in splenocytes of insulin 2-KO NOD mice to insulin 1 B:9-23 peptide was increased (P<0.05), suggesting that expanding insulin-reactive T cells may contribute to the acceleration of diabetes in insulin 2-KO mice. Based on those observations, we hypothesize that insulin 1 is a crucial T cell antigen in murine autoimmune diabetes and modification of anti-insulin autoimmunity can be applicable to antigen-based therapy for human type 1 diabetic patients.

Rituximab combined with a small dose of melphalan for a refractory follicular lymphoma patient.

A 48-year-old male patient with follicular lymphoma, grade II, stage IV, was treated with CHOP, ESHAP and MACOP-B, resulting in partial remission. After 9 months, the disease progressed and several chemotherapy agents, including three courses of rituximab combined with etoposide, sobuzoxane or methotrexate, only resulted in a stable disease response. However, the fourth course of rituximab combined with a small dose of melphalan produced excellent results and the complete response continued for more than 15 months. It is possible that these two drugs may act synergistically.