Variant spectrum of PIEZO1 and KCNN4 in Japanese patients with dehydrated hereditary stomatocytosis.

Hereditary stomatocytosis (HSt) is a type of congenital hemolytic anemia caused by abnormally increased cation permeability of erythrocyte membranes. Dehydrated HSt (DHSt) is the most common subtype of HSt and is diagnosed based on clinical and laboratory findings related to erythrocytes. PIEZO1 and KCNN4 have been recognized as causative genes, and many related variants have been reported. We analyzed the genomic background of 23 patients from 20 Japanese families suspected of having DHSt using a target capture sequence and identified pathogenic/likely pathogenic variants of PIEZO1 or KCNN4 in 12 families.

[Insulin-derived amyloidosis (insulin ball) and skin-related complications of insulin therapy].

Skin-related complications of insulin therapy have long been a problem as a factor interfering with insulin therapy. Among the traditional skin-related complications, lipoatrophy and insulin allergy have decreased markedly with the development of insulin preparations, but lipohypertrophy is still common in insulin-treated patients. Recently, there have been more reports of a skin-related complication called insulin-derived amyloidosis or insulin ball. Insulin-derived amyloidosis is a condition in which injected insulin becomes amyloid protein and is deposited at the injection site. Insulin-derived amyloidosis causes poor glycemic control and increased insulin dose requirements, which are caused by decreased insulin absorption. Lipohypertrophy also decreases insulin absorption, but insulin-derived amyloidosis causes a more significant decrease in insulin absorption and has a greater clinical impact. Therefore, it is important to make a differential diagnosis between insulin-derived amyloidosis and lipohypertrophy, but sometimes it is difficult to distinguish the two and imaging studies are required. The diagnosis of insulin-derived amyloidosis is often difficult in the general practice, and its pathogenesis and prevalence have not been fully clarified. Recently, it has been reported that insulin-derived amyloidosis can be toxic, suggesting an association with minocycline use. The treatment of insulin-derived amyloidosis and lipohypertrophy is to avoid the site of amyloidosis or lipohypertrophy and inject insulin, but the dose of insulin injection should be reduced. Prevention of both insulin-derived amyloidosis and lipohypertrophy is important, and for this purpose, observations of the insulin injection site and instruction on appropriate insulin injection techniques are necessary, and multidisciplinary cooperation is extremely important.

Multiple Endocrine Neoplasia Type 1 with Functional Parathyroid Cysts.

A 51-year-old woman was admitted because of hypercalcemia. Neck ultrasonography and computed tomography revealed the presence of parathyroid cysts on both sides. After primary hyperparathyroidism was diagnosed by technetium-99m-methoxyisobutylisonitrile scintigraphy, the patient was successfully treated with total parathyroidectomy and autotransplantation. She also had a non-functioning pancreatic neuroendocrine tumor, prolactinoma, and adrenal tumors with subclinical Cushing's syndrome. Given these clinical features and her family history, multiple endocrine neoplasia type 1 (MEN1) was suspected, and germline DNA sequencing revealed a missense mutation (c.1013T>C, [corrected] p.Leu338Pro) in exon 7 of MEN1. This case demonstrates the phenotypic and genetic diversity of MEN1.

A case of nonislet cell tumor hypoglycemia associated with malignant mesothelioma requiring a multifaceted approach for optimal glycemic control.

Nonislet tumor hypoglycemia (NICTH) is a rare paraneoplastic syndrome characterized by refractory hypoglycemia, which often requires multifaceted therapy. We reported a case of a patient with pleural malignant mesothelioma and developed NICTH, for which chemotherapy, glucocorticoids, and nutrition were given to achieve optimal glycemic control.

Regulation of Endothelium-Reticulum-Stress-Mediated Apoptotic Cell Death by a Polymethoxylated Flavone, Nobiletin, Through the Inhibition of Nuclear Translocation of Glyceraldehyde 3-Phosphate Dehydrogenase in Retinal Müller Cells.

In the early stages of diabetic retinopathy (DR), subtle biochemical and functional alterations occur in Müller cells, which are one of the components of the blood-retinal barrier (BRB). Müller cells are the principal glia of the retina and have shown a strong involvement in the maintenance of homeostasis and the development of retinal tissue. Their functional abnormalities and eventual loss have been correlated with a decrease in the tight junctions between endothelial cells and a consequent breakdown of the BRB, leading to the development of DR. We demonstrated that the endothelium reticulum (ER) triggers Müller cell death and that nuclear accumulation of glyceraldehyde 3-phosphate dehydrogenase is closely associated with ER-induced Müller cell death. In addition, induction of ER stress in Müller cells increased vascular endothelial growth factor expression but decreased pigment-epithelium-derived factor (PEDF) expression in Müller cells. We found that nobiletin, a polymethoxylated flavone from citrus explants, exerts protective action against ER-stress-induced Müller cell death. In addition, nobiletin was found to augment PEDF expression in Müller cells, which may lead to the protection of BRB integrity. These results suggest that nobiletin can be an attractive candidate for the protection of the BRB from breakdown in DR.

Insulin-derived amyloidosis without a palpable mass at the insulin injection site: A report of two cases.

To date, almost all case reports of insulin-derived amyloidosis described the presence of a subcutaneous mass that was observable on physical examination. This report presents two cases of insulin-derived amyloidosis without palpable masses at insulin injection sites. In both cases, blood glucose concentrations improved, and the insulin dose could be reduced by an average of 45% after changing the insulin injection sites. The insulin absorption at the site was reduced to at most 40% of that at a normal site in one case. Magnetic resonance imaging and ultrasonography were useful to screen and differentiate insulin-derived amyloidosis without a palpable mass. This report showed that insulin-derived amyloidosis without a palpable mass can be present at the insulin injection site, and has similar clinical effects to insulin-derived amyloidosis with palpable masses.

Toxicity of insulin-derived amyloidosis: a case report.

BACKGROUND: Insulin-derived amyloidosis is a skin-related complication of insulin therapy that interferes with insulin therapy. Although toxicities of in vitro-formed insulin amyloid fibrils have been well studied, the toxicity of insulin-derived amyloidosis remains to be clarified. CASE PRESENTATION: A 58-year-old man with type 2 diabetes mellitus underwent a lower limb amputation due to diabetic gangrene. Several antibiotics including minocycline were administered for infection and sepsis. A hard mass at the insulin injection sites in the lower abdomen was discovered by chance four months later. Although no abnormal findings in the surface skin of the mass were observed, necrotic tissue was seen around the mass when a biopsy was performed. Histological and toxicity studies were performed for this patient and four other patients with abdominal masses at insulin injection sites. Histological and immunohistochemical studies showed that the masses had typical characteristics of amyloid deposits in all cases, whereas necrotic findings were seen adjacent to the amyloid deposit only in the case presented. Toxicity studies indicated that the amyloid tissue from the present case had significant cell toxicity compared to the control skin tissue or the amyloid tissues from the other four cases. CONCLUSIONS: This report showed that toxic insulin-derived amyloidosis can occur. In addition, this report suggested that toxic insulin-derived amyloidosis may cause necrosis in the surrounding tissue. Although the toxic amyloid deposit of insulin-derived amyloidosis was found in only one patient, no structural differences between toxic and non-toxic deposits were seen on histological and immunohistochemical studies.

In vitro studies on nobiletin isolated from citrus plants and the bioactive metabolites, inhibitory action against gelatinase enzymatic activity and the molecular mechanisms in human retinal Müller cell line.

Diabetic retinopathy (DR) is the most common cause of vision loss in patients with diabetes mellitus. Despite the presence of effective therapy, DR is still a significant health burden. A recent research suggests that matrix metalloproteinases (MMPs) could be promising targets, which exert multiple actions on early- and late-stage pathogenesis of DR. Among the MMP family, gelatinases (MMP-2 and MMP-9) act as potent proinflammatory, proangiogenic, and pro-apoptotic factors. Therefore, the pharmacological inhibitory effect of gelatinases on retinal MMP-producing cells may be useful in the treatment or prevention of DR. Nobiletin isolated from citrus plants is a multi-functional polymethoxylated flavone, which exerts biological effects including inhibitory action against MMP activity in several cancer cells. In the present study, we demonstrated that nobiletin isolated from citrus plants attenuated MMP-9 enzymatic activity through the suppression of transcription for MMP-9 gene expression and augmentation of TIMP-1 production in retinal Müller cells. Nobiletin regulated MMP-9 gene expression and TIMP-1 by inhibiting the PI3K/Akt signaling pathway. In addition, we observed the augmentation of inhibitory action against MMP-9 enzymatic activity by 4'-demethylated nobiletin, which is a major metabolite of nobiletin. We believe that the enhancement of inhibitory action against MMP-9 enzymatic activity by 4'-demethylated nobiletin is through the dual inhibition on Erk1/2 and Akt phosphorylation. The structure-activity relationship analysis revealed that, for the enhancement of inhibitory action against MMP-9 enzymatic activity, demethylation at position 4' in B-ring was a key structural modification in Müller cells, which are an important source of MMPs found in vitreous fluid and retinal tissues in retinal proliferative diseases. These results suggested that nobiletin, derived from a natural source, may serve as a novel MMP inhibitor with minimal side effects, and lead compound for the design of more efficacious drugs.

Insulin-derived amyloidosis and poor glycemic control: a case series.

OBJECTIVES: Insulin-derived amyloidosis is a rare skin-related complication of insulin therapy. The purpose of this study was to show the effects of insulin-derived amyloidosis on blood glucose levels, insulin dose requirements, and insulin absorption. METHODS: Seven patients were found to have insulin-derived amyloidosis at the Tokyo Medical University Ibaraki Medical Center. The clinical characteristics and insulin therapy of the 7 patients were investigated. Insulin absorption was studied by comparing the serum insulin levels after insulin injections into insulin-derived amyloidosis sites versus injections into normal sites in 4 patients. RESULTS: When the insulin-derived amyloidosis was discovered, the mean hemoglobin A1c level was 9.3%, and the mean daily insulin dose was 57 units. After changing the injection sites to avoid the insulin-derived amyloidosis, the blood glucose concentrations improved, and the mean daily insulin dose could be reduced to 27 units (P = .035; 53% reduction). The insulin absorption at insulin-derived amyloidosis sites was 34% of that at normal sites (P = .030). CONCLUSIONS: Insulin-derived amyloidosis caused poor glycemic control and increased insulin dose requirements because of impairments in insulin absorption.

Cell-mediated contraction of vitreous explants from chicken embryo: possibility of screening for therapeutic agents against proliferative vitreoretinal diseases.

PURPOSE: We aimed to establish a novel screening system for identifying potential therapeutic agents for treating proliferative vitreoretinal diseases (PVDs). In this study, we focused on vitreous explants from chicken embryos and evaluated the usefulness of quantitatively analyzing the effects of potential candidates on cell-mediated vitreous contraction, which leads to blindness in PVDs. METHODS: Vitreous explants were extracted from 19-day-old embryonic chickens and then incubated with retinal Müller cells or endothelial cells to permit cell adhesion. After cell adhesion occurred, we examined the effect of the attached cells on the wet weight of vitreous explants as an index of vitreous contraction. We also performed hematoxylin and eosin staining to characterize the cell morphology on the vitreous surface. RESULTS: Contraction of the vitreous explants was observed after cell adhesion of not only retinal Müller cells but also endothelial cells. We confirmed the adhesion of these cells on vitreous explants and estimated the number of adherent cells with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) analysis. The cells on the vitreous surface presented an elongated fibroblast-like phenotype. Integrin was found to be a receptor involved in cell adhesion on the vitreous surface. DISCUSSION: Our results suggest that vitreous explants from chicken embryos may be novel useful tools for screening antiadhesion therapeutic agents in PVDs. This preliminary study must be validated with human vitreous and human retinal pigment epithelial cells.

Protein kinase C-mediated regulation of matrix metalloproteinase and tissue inhibitor of metalloproteinase production in a human retinal müller cells.

PURPOSE: Matrix metalloproteinases (MMPs) play an important role in the degradation of extracellular matrix (ECM) proteins in the retina. Breakdown of ECM proteins results in neovascularization and tractional retinal detachment, which eventually lead to the symptoms of proliferative diabetic retinopathy. Müller cells are reported to be one of the MMP-producing cells in the retina. However, the molecular mechanism of MMP production derived from Müller cells remains to be fully elucidated. MATERIALS AND METHODS: The human retinal Müller cell line (MIO-M1) was continuously-subcultured in high glucose (25 mM) condition. After the cells reached confluence, they were treated for 24 h with phorbol ester and/or a protein kinase C (PKC) inhibitor, GF109203X in high (25 mM) or low (5 mM) glucose condition. Gelatinase activities in conditioned medium were assessed using gelatin zymography. RT-PCR was performed to analyze the mRNA expression level of MMP-9. Western blot analysis used to detect the protein expression of tissue inhibitors of metalloproteinases (TIMPs). Electrophoresis mobility shift assay was conducted to examine transcription factors involved in MMP-9 transcription. RESULTS: We demonstrated the protein kinase C (PKC)-mediated regulation of proMMP-9 transcription and protein expression through the action of phorbol ester, an activator of PKC. Moreover, we demonstrated the expression of TIMPs, known as natural inhibitors of MMPs at the protein level in a human retinal Müller cell line for the first time, and report that production of these proteins was also regulated in a PKC-dependent manner. CONCLUSION: Our results suggest that imbalance between MMP and TIMP proteins may promote neovascularization by PKC activation in retinal Müller cells. In addition, the development of novel compounds with regulatory action on MMP and TIMP production through inhibiting PKC activity in retinal Müller cells may lead to new therapeutic approaches for the treatment and prevention of diabetic retinopathy.

Successful treatment of postsurgical hypoparathyroidism by intramuscular injection of vitamin D3 in a patient associated with malabsorption syndrome due to multiple abdominal surgeries.

A 56-year-old patient with postsurgical hypothyroidism and hypoparathyroidism associated with gastrointestinal malabsorption syndrome was prescribed with L: -thyroxine and 1alpha(OH)D(3) at a massive daily dosage of 600 and 39 mug, respectively. Although the patient became nearly euthyroid, she had been hypocalcemic, requiring frequent intravenous injection of calcium gluconate to prevent tetany. Because the serum level of 1,25(OH)(2)D hardly increased after an oral intake of 21 microg 1alpha(OH)D(3), vitamin D(3) was administered intramuscularly. After stoss therapy (600,000 IU), the patient has been receiving 300,000 IU vitamin D(3) at intervals of 2-4 months so that she remained slightly hypocalcemic (7-8 mg/dl). At 1.5 years later, serum levels of 25(OH)D and 1,25(OH)(2)D were maintained at about 60 ng/ml and 30-50 pg/ml, respectively, and renal function was maintained well. These data suggest that intramuscular injection of 300,000 IU vitamin D(3) at an interval of a few months to maintain a slightly increased serum level of 25(OH)D and a slightly decreased serum level of calcium is a safe and cost-effective treatment in such a parathyroid hormone-deficient hypoparathyroid patient with malabsorption syndrome.

A trial to assess the amount of insulin antibodies in diabetic patients by surface plasmon resonance.

OBJECTIVE: To measure the amount and affinity of insulin antibodies, we performed a trial to establish a new method for quantitative and qualitative analysis of these antibodies by using surface plasmon resonance (BIAcore system). METHODS: Real-time detection of insulin antibody interaction and kinetic analysis were performed using the BIAcore system. PATIENTS OR MATERIALS: Eight diabetic patients with insulin antibodies and whose fasting total immunoreactive insulin levels were more than 100 microU/ml were selected. The patients with and without recurrent hypoglycemia were classified into hypoglycemic episode-positive or hypoglycemic episode-negative groups, respectively. Seven diabetic patients without insulin antibodies were selected as controls. RESULTS: In the 8 patients, the concentration of insulin antibodies ranged from 2.91 to 16.3 microg/ml and insulin antibodies were not detected in the control group. The apparent KD (dissociation constant) and kd (the dissociation rate constant) values of the patients were much larger than those seen for the anti-human insulin monoclonal antibody. The KD values were significantly higher in the hypoglycemic episode-positive group than in the hypoglycemic episode-negative group (p<0.05). No significant differences in the concentration, the ka (the association rate constant) and the kd values were noted between the groups. CONCLUSION: The data suggests that insulin antibodies of the patients have an apparently lower affinity status in sera as compared with that for the anti-human insulin monoclonal antibody, and dissociate easily from the immune-complex in the sera, especially in cases where there is recurrent hypoglycemia in the patients. Therefore insulin antibody characteristics are one of the causative factors in hypoglycemic episodes.

A study of urinary myo-inositol as a sensitive marker of glucose intolerance.

BACKGROUND: We assessed the possibility of using myo-inositol as a marker of glucose intolerance. METHODS: We measured urinary myo-inositol enzymatically before and 2 h after a 75-g oral glucose tolerance test in 564 volunteers, who were divided into four groups [normal glucose tolerance (NGT), impaired fasting glucose (IFG), impaired glucose tolerance (IGT), and diabetes mellitus (DM)]. Furthermore, we classified NGT into NGT-A (2-h blood glucose <120 mg/dl and 2-h glucosuria <50 mg/dl) and NGT-B (remaining NGT subjects). We then compared deltamyo-inositol (myo-inositol/creatinine ratio: 2-h after glucose load--before load) of each group to investigate the relationship between glucose intolerance and deltamyo-inositol. RESULTS: The glucose tolerance of NGT-B appeared to have deteriorated compared with NGT-A as determined by blood glucose, insulin, and glucosuria. There was very little effect of gender or age on deltamyo-inositol in NGT-A. deltamyo-inositol was significantly higher than that in NGT-A (0.5+/-7.1 mg/g Cr) not only in IFG (8.7+/-19.5 mg/g Cr, P<0.0001), IGT (14.8+/-22.9 mg/g Cr, P<0.0001) and DM (79.5+/-37.1 mg/g Cr, P<0.0001), but in NGT-B (7.4+/-12.7 mg/g Cr, P<0.0001). With 2 mg/g Cr as a tentative cut-off for deltamyo-inositol to detect NGT-A, sensitivity and specificity were 68% and 72%, respectively. CONCLUSIONS: The deltamyo-inositol can be use of a non-invasive and sensitive marker for glucose intolerance.