Identification of a New Enhancer That Promotes Sox9 Expression by a Comparative Analysis of Mouse and Sry-Deficient Amami Spiny Rat.

INTRODUCTION: Testis differentiation is initiated by the SRY gene on the Y chromosome in mammalian species. However, the Amami spiny rat, Tokudaia osimensis, lacks both the Y chromosome and the Sry gene and acquired a unique Sox9 regulatory mechanism via a male-specific duplication upstream of Sox9, without Sry. In general mammalian species, the SRY protein binds to a testis-specific enhancer to promote SOX9 gene expression. Several enhancers located upstream of Sox9/SOX9 have been reported in mice and humans. In particular, the binding of SRY to the highly conserved enhancer Enh13 is thought to be a common mechanism underlying testis differentiation and sex determination in mammals. METHODS: Sequences of T. osimensis homologues of three Sox9 enhancers that were previously reported in mice, Enh8, Enh14, and Enh13, were determined. We performed in vitro assays to confirm enhancer activity involved in Sox9 regulation in T. osimensis. RESULTS: T. osimensis Enh13 showed enhancer activity when co-transfected with NR5A1 and SOX9. Mouse Enh13 was activated by NR5A1 and SRY; however, T. osimensis Enh13 did not respond to SRY, even though the binding sites of SRY and NR5A1 were conserved. To identify the key sequence that is present in mouse but absent from T. osimensis, we performed reporter gene assays using vectors in which partial sequences of T. osimensis Enh13 were replaced with mouse sequences. For T. osimensis Enh13 in which the second half (approximately 430 bp) was replaced with the corresponding mouse sequence, activity in response to NR5A1 and SRY was recovered. Further, reporter assays revealed that multiple regions in the second half of the mouse Enh13 sequence are required for the response to NR5A1 and SRY. The latter 49 bp was particularly important and contained four binding sites for three transcription factors, POU2F1, HOXA3, and GATA1. CONCLUSION: We showed that there are unknown sequences responsible for the interaction between NR5A1 and SRY and mEnh13 based on comparative analyses of Sry-dependent and Sry-independent species. Our comparative analyses revealed new molecular mechanisms underlying mammalian sex determination.

Late recurrence in patients with non-muscle-invasive bladder cancer after 5-year cancer-free periods.

OBJECTIVES: There is no consensus about the follow-up schedule after 5-year cancer-free periods. In this study, we aimed to elucidate the risk factors for the recurrence in patients with non-muscle-invasive bladder cancer who remained cancer free for more than 5 years. METHODS: Data from six Japanese institutions were retrospectively reviewed. Among the patients with non-muscle-invasive bladder cancer who were treated with transurethral resection of bladder tumor between 1990 and 2013, those who had no recurrence for more than 5 years were included in this study. The Kaplan-Meier method and Cox hazards model were used to estimate recurrence-free survival and to determine the pathologic and clinical factors affecting late recurrence. RESULTS: In total, 434 patients were enrolled in this study. Of these patients, 55 patients (12.7%) experienced late recurrence. The median follow-up time was 8.9 years (interquartile range 6.9-11.3 years). Prior history of bladder cancer before the most recent transurethral resection was a significant predictor for late recurrence (hazard ratio 1.99 [95% confidence interval 1.13-3.47], P = 0.019), although other clinical factors including tumor grade, pathologic stage, tumor multiplicity, and current risk classification systems were not associated with late recurrence. CONCLUSIONS: Late recurrence after a long tumor-free period is not rare and it was not predicted by current risk classification systems. Only prior history of bladder cancer was a significant predictor for late recurrence in this study.

What are the Optimal Renal Ultrasound Parameters for Detecting Small Kidney in Young Children?

INTRODUCTION: Recent guidelines do not recommend routine screening of vesicoureteral reflux after a first febrile urinary tract infection in children without abnormal findings on ultrasound or atypical/recurrent urinary tract infection. Currently, there are no clear ultrasonographic parameters for detecting abnormalities in renal size, especially in young children. The aim of the present study was to determine an optimal cutoff value for detecting small kidney in children without apparent congenital anomalies except vesicoureteral reflux by retrospective chart review. PATIENTS AND METHODS: Children aged ≤3 years who had undergone nuclear renal scans and ultrasound were enrolled. Small kidney was defined as split renal function of <40%. Optimal cutoff values of various ultrasonographic parameters for detecting small kidney were calculated. RESULTS: Of the 69 children included in the present study, small kidney was identified in 20. There was a significant difference in renal size between each kidney in patients with small kidney, whereas there was no significant difference in those without small kidney. With a ratio of estimated renal area of 74.26%, maximum area under the curve with the highest sensitivity, specificity, positive predictive value, negative predictive value, and accuracy rate were obtained. In addition, simple measurement of renal length with a cutoff of 4.97 cm showed high specificity comparable with estimated renal area. CONCLUSION: Small kidney may be screened by two-dimensional measurement on ultrasonographic examination, even in young children. With the cutoff described, risk stratification or an individualized approach may be possible.

Comparison of the effects of three kinds of glucose-lowering drugs on non-alcoholic fatty liver disease in patients with type 2 diabetes: A randomized, open-label, three-arm, active control study.

AIMS/INTRODUCTION: Non-alcoholic fatty liver disease (NAFLD) is often observed in individuals with type 2 diabetes mellitus, and it is known that the presence of type 2 diabetes mellitus leads to the aggravation of NAFLD. The aim of this study was to compare the possible effects of three kinds of oral hypoglycemic agents on NAFLD in individuals with type 2 diabetes mellitus. MATERIALS AND METHODS: We carried out a prospective clinical trial (a randomized and open-label study) in patients with type 2 diabetes mellitus and NAFLD. A total of 98 patients were randomly allocated either to the dapagliflozin (n = 32), pioglitazone (n = 33) or glimepiride (n = 33) group, and the patients took these drugs for 28 weeks. The primary end-point was the change of the liver-to-spleen ratio on abdominal computed tomography. RESULTS: There was no difference in baseline clinical characteristics among the three groups. Dapagliflozin, pioglitazone and glimepiride ameliorated hyperglycemia similarly. Bodyweight and visceral fat area were significantly decreased only in the dapagliflozin group. Serum adiponectin levels were markedly increased in the pioglitazone group compared with the other two groups. Dapagliflozin and pioglitazone, but not glimepiride, significantly increased the liver-to-spleen ratio, and the effects of dapagliflozin and pioglitazone on the liver-to-spleen ratio were comparable. CONCLUSIONS: The present study showed that the decrease of visceral fat area and the increase of adiponectin level contributed to the improvement of NAFLD in patients with type 2 diabetes mellitus. Furthermore, dapagliflozin and pioglitazone exerted equivalent beneficial effects on NAFLD in patients with type 2 diabetes mellitus, although it seemed that these two drugs had different mechanisms of action.

[PUBERTAL TESTICULAR TORSION OF POLYORCHIDISM DIAGNOSED IN INFANCY: A CASE REPORT].

Left polyorchidism was found in a 2-month-old boy with a left scrotal mass. As he was asymptomatic and all testes were in the scrotum, he was conservatively followed up. At 17 years of age, he presented with left acute scrotum due to testicular torsion of the left supernumerary testis. Counterclockwise 720-degree rotation of the left supernumerary testis was noted during emergency surgery, and orchidopexy of the 3 testes (2 left testes and 1 right testis) was performed. Biopsy of the left supernumerary testis demonstrated spermatogenesis and no malignancy. One and a half years after surgery, all testes were viable without atrophy.Polyorchidism is a rare condition and there is no consensus on the management of asymptomatic cases detected early in life. The position of the supernumerary testis (intrascrotal or extrascrotal) is important when deciding the management strategy because of the risk of malignancy. If conservative management is selected initially, elective surgery, such as prophylactic orchiectomy or orchidopexy, may be needed because of the risk of malignancy and torsion.

Long-Term Treatment with Thrombomodulin Improves Functional Outcomes after Cerebral Ischemia Even if Administration is Delayed.

Our previous study indicated that recombinant human soluble thrombomodulin (rhsTM) could attenuate brain damage when administered as a bolus in the cerebral ischaemic early phase. Then, we considered that treatment with rhsTM may show therapeutic effects even when administered in the ischaemic delayed phase, because rhsTM has an action of inhibiting high-mobility group box 1 (HMGB1) as a late mediator of lethal systemic inflammation. This study was performed to investigate the effects of delayed treatment with rhsTM on ischaemic brain damage induced by high HMGB1 level in mice subjected to 4-hour middle cerebral artery occlusion (MCAO). One day after MCAO, rhsTM was administered intraperitoneally at a dose of 1 or 5 mg/kg once a day for 7 days. Neurological score, motor coordination and HMGB1 levels were measured 1, 3 and 7 days after MCAO. The presence of activated microglia was evaluated 7 days after MCAO. Systemic HMGB1 levels increased 1 to 7 days after MCAO and were higher at 7 days compared with day 1. At the same time, survival rate decreased, and activated microglia increased in the infarct area. Treatment with rhsTM improved neurological score, motor coordination, survival and prevented brain damage. Moreover, rhsTM decreased both HMGB1 level and number of activated M1 microglia. The results of this study indicated that rhsTM improved functional outcomes via inhibition of HMGB1 up-regulation and M1 microglial activation in the cerebral ischaemic delayed phase. rhsTM may become a new therapeutic agent with a wide therapeutic time window in patients with cerebral ischaemia.

There is a Close Association Between the Recovery of Liver Injury and Glycemic Control after SGLT2 Inhibitor Treatment in Japanese Subjects with Type 2 Diabetes: A Retrospective Clinical Study.

INTRODUCTION: Sodium-glucose co-transporter 2 (SGLT2) inhibitors function not only to reduce hyperglycemia but also to ameliorate liver injury and reduce body weight. The aim of this study was to examine in which subjects SGLT2 inhibitors are more effective for glycemic control, liver injury, and obesity in Japanese subjects with type 2 diabetes mellitus. METHODS: We enrolled a total of 156 subjects with type 2 diabetes who initiated SGLT2 inhibitor treatment after September 1, 2014 in Kawasaki Medical School (Protocol No. 2375). We evaluated the alteration of glycemic control, liver injury, body mass composition, and various clinical parameters. RESULTS: SGLT2 inhibitors significantly ameliorated glycemic control and improved liver injury in Japanese subjects with type 2 diabetes. SGLT2 inhibitors were more effective for liver injury when glycemic control was improved with SGLT2 inhibitors. In multivariate analyses, the amelioration of glycemic control was an independent determinant factor for the improvement of liver damage in Japanese subjects with type 2 diabetes. The reverse was also correct; the improvement of liver damage was an independent determinant factor for the amelioration of glycemic control. CONCLUSION: Recovery of liver injury with SGLT2 inhibitor treatment was closely associated with their effects on glycemic control in Japanese subjects with type 2 diabetes.

Werner Syndrome and Diabetes Mellitus Accompanied by Adrenal Cortex Cancer.

Werner syndrome is a rare genetic disease characterized by progeria, diabetes mellitus, cataracts and various types of malignancy. However, there are few reports showing adrenal cortex cancer in subjects with Werner syndrome. We herein report an extremely rare case of Werner syndrome accompanied by adrenal cortex cancer. Based on the data obtained from blood samples, computed tomography, magnetic resonance imaging and 131I adosterol scintigraphy, we diagnosed this subject with adrenal cortex cancer and Cushing's syndrome. Since the prognosis of adrenal cancer is very poor, we should be aware of the possibility of adrenal cancer occurring in subjects with Werner syndrome.

Hypoglycemic Coma Induced by the Use of Succinic Acid Cibenzoline in Frail Late-stage Elderly Subjects.

Succinic acid cibenzoline (CZ) is an antiarrhythmic agent often used for the treatment of tachyarrhythmia. However, hypoglycemia should be avoided in the treatment of diabetes. We herein report two late-stage elderly subjects who experienced a severe and prolonged hypoglycemic coma after the usage of CZ. These cases suggest that, when CZ is administered to elderly subjects with renal dysfunction and/or frailty, we should be aware of the possibility that this medicine may induce hypoglycemia and should adjust the dose as appropriate and monitor the concentration of CZ to avoid severe hypoglycemia.

Dramatic mitigation of bone pain after phosphorus replacement therapy in a subject with FGF23-related hypophosphatemic osteomalacia.

INTRODUCTION: Fibroblast growth factor 23 (FGF23) is secreted from bone and suppresses the absorption of phosphorus in renal proximal tubule and in intestinal tract. Therefore, the increase of serum FGF23 levels leads to hypophosphatemic situations. Tumor-induced osteomalacia is often induced by various tumors, but it is often difficult to identify the localization of tumor, because most of the FGF23-producing tumors are small and could be observed in any part of the body. CASE DESCRIPTION: Here we report a case of elderly female subject with FGF23-related hypophosphatemic osteomalacia who repeatedly experienced severe bone pain and fragility fracture in various parts of the body. Although we failed to identify the localization of tumor in this subject even with various examination, after starting phosphorus replacement therapy with relatively small amounts of calcium phosphate (1.5 g/day) (phosphorus content: 270 mg), hypophosphatemia was ameliorated and repeated bone pain was dramatically mitigated without any surgical operation. DISCUSSION AND EVALUATION: Even when we fail to identify the localization of tumor in subjects with FGF23-related hypophosphatemic osteomalacia, phosphorus replacement therapy for hypophosphatemia could reduce the bone pain. CONCLUSIONS: We should be aware of the possibility that phosphorus replacement therapy exert marked beneficial effects for the reduction of bone pain in subjects with FGF23-related hypophosphatemic osteomalacia even when we fail to identify tumor localization.

Association of GA/HbA1c ratio and cognitive impairment in subjects with type 2 diabetes mellitus.

AIMS: The aim of this study was to search for factors influencing cognitive impairment and to clarify the association between the fluctuation of blood glucose levels and cognitive impairment in elderly Japanese subjects with type 2 diabetes. METHODS: We recruited 88 relatively elderly subjects (≥65years old) with type 2 diabetes who were hospitalized in Kawasaki Medical School from January 2014 to December 2015. We evaluated the fluctuation of blood glucose levels with glycoalbumin (GA)/hemoglobin A1c (HbA1c) ratio, and estimated cognitive impairment with Hasegawa dementia scale-revised (HDS-R) score and mini mental state examination (MMSE) score. RESULTS: Multivariate analyses showed that GA/HbA1c ratio and urinary albumin excretion, but not hypoglycemia, were independent determinant factors for cognitive impairment in elderly Japanese subjects with type 2 diabetes. CONCLUSIONS: The fluctuation of blood glucose levels per se is closely associated with cognitive impairment in elderly subjects with type 2 diabetes even when hypoglycemia is not accompanied. Since it is very easy to calculate GA/HbA1c ratio, we should check this ratio so that we can reduce the fluctuation of blood glucose levels especially in elderly subjects with type 2 diabetes.

Case of disseminated pyomyositis in poorly controlled type 2 diabetes mellitus with diabetic ketoacidosis.

Primary pyomyositis is a pyogenic and uncommon infection of skeletal muscle, which is mainly observed in tropical areas and/or human immunodeficiency virus patients. In non-human immunodeficiency virus infected patients, the most common cause is diabetes mellitus. Because of its rarity, the accurate diagnosis is often challenging. Staphylococcus aureus is the most common causative bacteria. According to the severity, pyomyositis is divided into three stages, and the late stage is occasionally lethal. The present case was compatible with the most advanced stage. Therefore, it was very difficult to save her life without precise and timely diagnosis. Furthermore, in the invasive stage, surgical drainage and broad-spectrum antibiotics should be given for a long enough period. Here, we report a case of a Japanese woman who developed disseminated abscesses under poorly controlled diabetic conditions accompanied by ketoacidosis, but was successfully treated without any sequelae.

Rapid onset of syndrome of inappropriate antidiuretic hormone secretion induced by duloxetine in an elderly type 2 diabetic patient with painful diabetic neuropathy.

Diabetic neuropathy is the most common diabetic complication. Duloxetine, a serotonin noradrenaline reuptake inhibitor (SNRI), is widely used for the treatment of diabetic painful neuropathy (DPN) because of the efficacy and safety profile. Syndrome of inappropriate antidiuretic hormone secretion, which is strongly associated duloxetine, is a rare but occasionally life-threatening adverse effect. Here, we report a case of syndrome of inappropriate antidiuretic hormone secretion that rapidly developed after starting duloxetine in an elderly Japanese female type 2 diabetes mellitus patient. Furthermore, we discuss the possible relationship between the onset of syndrome of inappropriate antidiuretic hormone secretion and the gene polymorphism of cytochrome P450 isoform 1A2 and 2D6, both of which are responsible for duloxetine metabolism.