Establishment of sandwich ELISA for soluble alpha-Klotho measurement: Age-dependent change of soluble alpha-Klotho levels in healthy subjects

Background α-Klotho (αKl) regulates mineral metabolism such as calcium ion (Ca2+) and inorganic phosphate (Pi) in circulation. Defects in mice result in clinical features resembling disorders found in human aging. Although the importance of transmembrane-type αKl has been demonstrated, less is known regarding the physiological importance of soluble-type αKl (sαKl) in circulation. Objectives The aims of this study were: (1) to establish a sandwich ELISA system enabling detection of circulating serum sαKl, and (2) to determine reference values for sαKl serum levels and relationship to indices of renal function, mineral metabolism, age and sex in healthy subjects. Results We successively developed an ELISA to measure serum sαKl in healthy volunteers (n = 142, males 66) of ages (61.1 ± 18.5 year). The levels (mean ± SD) in these healthy control adults were as follows: total calcium (Ca; 9.46 ± 0.41 mg/dL), Pi (3.63 ± 0.51 mg/dL), blood urea nitrogen (BUN; 15.7 ± 4.3 mg/dL), creatinine (Cre; 0.69 ± 0.14 mg/dL), 1,25 dihydroxyvitamin D (1,25(OH)2D; 54.8 ± 17.7 pg/mL), intact parathyroid hormone (iPTH; 49.2 ± 20.6 pg/mL), calcitonin (26.0 ± 12.3 pg/mL) and intact fibroblast growth factor (FGF23; 43.8 ± 17.6 pg/mL). Serum levels of sαKl ranged from 239 to 1266 pg/mL (mean ± SD; 562 ± 146 pg/mL) in normal adults. Although sαKl levels were not modified by gender or indices of mineral metabolism, sαKl levels were inversely related to Cre and age. However, sαKl levels in normal children (n = 39, males 23, mean ± SD; 7.1 ± 4.8 years) were significantly higher (mean ± SD; 952 ± 282 pg/mL) than those in adults (mean ± SD; 562 ± 146, P < 0.001). A multivariate linear regression analysis including children and adults in this study demonstrated that sαKl correlated negatively with age and Ca, and positively with Pi. Finally, we measured a serum sαKl from a patient with severe tumoral calcinosis derived from a homozygous missense mutation of α-klotho gene. In this patient, sαKl level was notably lower than those of age-matched controls. Conclusion We established a detection system to measure human serum sαKl for the first time. Age, Ca and Pi seem to influence serum sαKl levels in a normal population. This detection system should be an excellent tool for investigating sαKl functions in mineral metabolism

Forebrain Ptf1a Is Required for Sexual Differentiation of the Brain

The mammalian brain undergoes sexual differentiation by gonadal hormones during the perinatal critical period. However, the machinery at earlier stages has not been well studied. We found that Ptf1a is expressed in certain neuroepithelial cells and immature neurons around the third ventricle that give rise to various neurons in several hypothalamic nuclei. We show that conditional Ptf1a-deficient mice (Ptf1a cKO) exhibit abnormalities in sex-biased behaviors and reproductive organs in both sexes. Gonadal hormone administration to gonadectomized animals revealed that the abnormal behavior is caused by disorganized sexual development of the knockout brain. Accordingly, expression of sex-biased genes was severely altered in the cKO hypothalamus. In particular, Kiss1, important for sexual differentiation of the brain, was drastically reduced in the cKO hypothalamus, which may contribute to the observed phenotypes in the Ptf1a cKO. These findings suggest that forebrain Ptf1a is one of the earliest regulators for sexual differentiation of the brain

Mucin-hypersecreting bile duct neoplasm characterized by clinicopathological resemblance to intraductal papillary mucinous neoplasm (IPMN) of the pancreas

<p>Abstract</p> <p>Background</p> <p>Although intraductal papillary mucinous neoplasm (IPMN) of the pancreas is acceptable as a distinct disease entity, the concept of mucin-secreting biliary tumors has not been fully established.</p> <p>Case presentation</p> <p>We describe herein a case of mucin secreting biliary neoplasm. Imaging revealed a cystic lesion 2 cm in diameter at the left lateral segment of the liver. Duodenal endoscopy revealed mucin secretion through an enlarged papilla of Vater. On the cholangiogram, the cystic lesion communicated with bile duct, and large filling defects caused by mucin were observed in the dilated common bile duct. This lesion was diagnosed as a mucin-secreting bile duct tumor. Left and caudate lobectomy of the liver with extrahepatic bile duct resection and reconstruction was performed according to the possibility of the tumor's malignant behavior. Histological examination of the specimen revealed biliary cystic wall was covered by micropapillary neoplastic epithelium with mucin secretion lacking stromal invasion nor ovarian-like stroma. The patient has remained well with no evidence of recurrence for 38 months since her operation.</p> <p>Conclusion</p> <p>It is only recently that the term "intraductal papillary mucinous neoplasm (IPMN)," which is accepted as a distinct disease entity of the pancreas, has begun to be used for mucin-secreting bile duct tumor. This case also seemed to be intraductal papillary neoplasm with prominent cystic dilatation of the bile duct.</p

当院における便潜血陽性者に対する大腸CT(CTコロノグラフィー)検査の有用性:大腸がん検診への導入と課題

大腸がん検診におけるスクリーニング検査としての大腸CT(CT colonography: CTC)検査の有用性を検討するために,当院における便潜血陽性者に対するCTCと大腸内視鏡検査の精度比較を行った.2009年7月から2014年1月までに川崎医科大学附属病院で施行されたCTC検査673件中,スクリーニング目的で行われた411件の中で便潜血陽性者に対して行われた183名を対象とした.全例CTC検査と同日に全大腸内視鏡検査も行った.対象とする病変は内視鏡観察あるいは病理組織学的に腺腫,がんと診断されたものとした.CTCの前処置は,経口腸管洗浄剤に水溶性造影剤による標識(タギング)を付けて行った.CT装置は16列Multi-slice CT(MSCT),腸管拡張は自動炭酸ガス注入器を使用した.CTC読影は,まず仮想内視鏡(3D)で行い,後に多断面再構成像(Multi-planar reconstruction: MPR 像(2D))を行う3D primary 法で行った.183名(男性98名,女性85名,年齢40～86歳,平均年齢62.1歳±0.8歳)のうち,病変を認めなかったのは87名(47.5%)であり,病変を認めたのは96名(53%)であった.総病変数は191個であり,うち6mm以上の病変は77個(40%)で,そのうち10mm以上のものは46個(24%)であった.大腸癌は25例(全病変中13%)で,うち腺腫内癌16例(全病変中8%)であった.側方発育型腫瘍は8例(4%)(大きさ平均17mm)であった.病変のうち,内視鏡的切除が行われたものは34病変であり,手術が行われたものは22病変であった.病変形態別による描出率は隆起型病変80%で,平坦型病変65%であった.病変サイズ別の精度は10mm以上の病変(n=46)で感度96%,陽性適中率98%であり,6mm以上の病変(n=77)で感度83%,陽性適中率79%であった.CTCは便潜血陽性者において良好な精度を示し,大腸がんスクリーニング法としての可能性がある.The purpose of this study was to estimate the sensitivity and specificity of CT colonography (CTC) for colorectal cancer screeing following positive fecal occult blood test (FOBT) in Japan. To compare detection rates of colorectal cancer and adenoma between CTC and optical total colonoscopy (TCS). This study included 183 patients with positive result of FOBT in Japanese colorectal cancer screening program. The patients had both CTC and TCS on the same day. 96 patients (53%) had colorectal lesions, on the other hand 87 patients had no lesions. The total number of lesions was 191, including 77 lesions 6 mm in maximum diameter and larger, including 46 lesions 10 mm and larger

Antibacterial-Agent-Immobilized Gelatin Hydrogel as a 3D Scaffold for Natural and Bioengineered Tissues

Hydrogels and their medical applications in tissue engineering have been widely studied due to their three-dimensional network structure, biocompatibility, and cell adhesion. However, the development of an artificial bile duct to replace the recipient&rsquo;s tissue is still desired. Some challenges remain in the tissue engineering field, such as infection due to residual artifacts. In other words, at present, there are no established technologies for bile duct reconstruction as strength and biocompatibility problems. Therefore, this study investigated hydrogel as an artificial bile duct base material that can replace tissue without any risk of infectious diseases. First, an antibacterial agent (ABA), Finibax (an ABA used for the clinical treatment of biliary tract infection), was immobilized in gelatin using a crosslinking agent, and the antibacterial properties of the gel and its sustainability were tested. Furthermore, the immobilized amount and the improvement of the proliferation of the human umbilical vein endothelial cells (HUVECs) were cultured as the ABA-Gelatin hydrogel was introduced to prepare a 3D scaffold. Finally, we performed hematoxylin and eosin (H&amp;E) staining after subcutaneous implantation in the rat. Overall, the ABA-Gelatin hydrogel was found to be viable for use in hydrogel applications for tissue engineering due to its good bactericidal ability, cell adhesion, and proliferation, as well as having no cytotoxicity to cells