Myogenic and adipogenic properties of goat skeletal muscle stem cells

The aims of the present study were to establish a culture system for goat skeletal muscle stem cells and to examine their myogenic and adipogenic properties in vitro. Cells were isolated from the skeletal muscle of the Shiba goat and cultured in vitro. Most of the cells were positive for myogenic markers, such as Pax7, MyoD, and desmin, and immunocytochemistry revealed they differentiated to form myotubes expressing myosin heavy chain, indicating they were highly myogenic. Myogenic differentiation was strongly suppressed by the addition of basic fibroblast growth factor, while proliferation was unaffected. When the cells were cultured in adipogenic differentiation medium, some of the cells differentiated into mature adipocytes that stained with Oil Red-O. These cells were immunocytochemically positive for adipogenic markers, including peroxisome proliferator-activated receptor-gamma (PPAR gamma) and CCAAT/enhancer-binding protein-alpha (C/EBP alpha). These results clearly demonstrate the presence of both myogenic and adipogenic stem cells in goat skeletal muscle.     

The Effects of GABA on embryonic gonadotropin-releasing hormone neurons in rat hypothalamic primary culture

Gonadotropin-releasing hormone (GnRH) neurons arise in the olfactory placode, migrate into the preoptic area (POA), and then extend axons to the median eminence during embryogenesis. Little information is available concerning the properties of GnRH neurons during the late gestational period when GnRH neurons reach the POA and form neuronal networks, although many studies have examined such properties during earlier developmental stages or the postnatal period. The present study was performed to elucidate the involvement of gamma-aminobutyric acid (GABA), one of the major neurotransmitters modifying GnRH neural activity, in regulation of GnRH gene expression on embryonic day 18.5 (E18.5) using transgenic rats expressing enhanced green fluorescence protein (EGFP) under the control of GnRH promoter. First, using RT-PCR, the mRNA of two isoforms of the GABA-synthesizing enzyme glutamic acid decarboxylase (GAD), GAD65 and GAD67 was detected in E18.5 embryonic POA-containing tissues. GAD67-positive cells were also demonstrated in close vicinity to GnRH-positive cells by immunohistochemistry, and immunoreactivity for both the GABA-A and GABA-B receptor subunits was detected in GnRH neurons. Next, primary cultures derived from anterior hypothalamic tissue of E18.5 embryos were prepared, and the effects of GABA and its agonists on GnRH promoter activity were evaluated using EGFP expression as a marker. GABA and the GABA-A receptor agonist muscimol, but not the GABA-B receptor agonist baclofen, significantly increased the EGFP-positive/GnRH-positive cell ratio. These results suggest that GABA plays a role in stimulating GnRH gene expression through GABA-A receptors in embryonic GnRH neurons in late gestational stages.     

CONTRAST‐ENHANCED ULTRASONOGRAPHIC FINDINGS IN THREE DOGS WITH PANCREATIC INSULINOMA

Abdominal ultrasonography is one of the most common diagnostic imaging modalities used for dogs with suspected insulinoma; however, pancreatic masses are clearly identified in fewer than half of affected dogs and benign pancreatic nodules can be difficult to differentiate from malignant ones. The purpose of this prospective study was to describe contrast‐enhanced ultrasonography (CEUS) characteristics of confirmed pancreatic insulinoma in a group of dogs. Inclusion criteria were as follows: (1) repeated hypoglycemia (blood glucose levels <60 mg/dl, twice or more); (2) elevated blood insulin levels with hypoglycemia; (3) pancreatic nodules detected with conventional ultrasonography; and (4) histological confirmation of pancreatic islet cell carcinoma. Immediately following conventional ultrasonography of the entire abdomen, CEUS of the pancreatic nodule and adjacent parenchyma was performed using contrast‐specific technology pulse inversion imaging and perflubutane microbubble contrast agent. Three dogs met inclusion criteria. Pancreatic nodules in all the three dogs became more clearly demarcated after injection of the contrast agent. Each nodule showed different enhancement patterns: markedly hyperechoic for 5 s, slightly hyperechoic for 1 s, and clearly hypoechoic for over 30 s. These results were not in complete agreement with previously reported CEUS findings in human patients with insulinoma. All nodules were surgically resected and histopathologically confirmed as malignant insulinomas. Findings from the current study indicated that contrast‐enhanced ultrasound may help to increase conspicuity of pancreatic insulinomas in dogs and that enhancement characteristics may be more variable in dogs than in humans.     

Improved survival and growth performances with photoperiod and feeding schedule manipulation in bagrid catfish Mystus nemurus (Cuvier & Valenciennes 1840) larvae

Manipulation of photoperiod: 24 h light (24L), 12 h light:12 h dark (12L:12D) and 24 h dark (24D); and feeding schedules: day and night feeding (DNF), day feeding (DF) and night feeding (NF) was conducted to determine effects on survival, cannibalism and growth of larval bagrid catfish Mystus nemurus 2–14 days after hatching (dAH). Photoperiod insignificantly affected all parameters. Feeding schedule significantly affected survival and total length at 6 (P < 0.049; P < 0.009), 10 (P < 0.033; P < 0.000) and 14 dAH (P < 0.013; P < 0.000), respectively, but affected cannibalism at 10 (P < 0.043) and 14 dAH (P < 0.013). Survival for DNF was significantly higher than DF. Cannibalism for DNF was significantly lower than NF at 10 and 14 dAH. Total length for DNF was significantly higher than DF and NF at 10 and 14 dAH. At 14 dAH, feeding schedule significantly affected feed intake, final weight and coefficient of variation. For feed intake and final weight, DNF was significantly higher than DF and NF. For coefficient of variation, NF was significantly higher than DF. This study suggests that larval bagrid catfish can be reared at 24L, 12L:12D or 24D but should be fed day and night for improved growth, survival and reduced cannibalism.     

Living Mulch with White Clover Improves Phosphorus Nutrition of Maize of Early Growth Stage

We examined the effect of a living mulch with white clover on the growth of maize in an Andisol. Maize was grown using a living mulch without fertilizer application, or by conventional cultivation with or without fertilizer application. Although the living mulch did not affect the amount of available phosphate in soil, the phosphorus concentration of maize shoots increased due to the living mulch compared with conventional cultivation without fertilizer application. Arbuscular mycorrhizal (AM) colonization of maize roots was higher with the living mulch than with conventional cultivation. These results suggested that a living mulch with white clover enhanced the phosphorus uptake by maize through AM colonization.     

Analysis of the mineral composition of taro for determination of geographic origin

The mineral composition of taro ( Colocasia esculenta (L.) Schott) was analyzed to develop a method to distinguish taro produced in Japan and China. The concentrations of 15 elements (Al, Ca, Cl, Mg, Mn, Br, Co, Cr, Cs, Fe, K, Na, Rb, Sc, Zn) were assayed using instrumental neutron activation analysis. The concentrations of NO(3)(-), SO(4)(2-), H(2)PO(4)(-), Cl(-), malate, and oxalate were measured by ion chromatography. The mean concentrations of H(2)PO(4)(-), Co, Cr, and Na significantly differed (P < 0.01) between taro grown in Japan and that grown in China. Discriminant analysis was performed to identify the most efficient combination of elements and compounds to discriminate the taro geographic origin. The highest percentage of correct classification was achieved with a two-variable model including H(2)PO(4)(-) and Co (100% for Japanese, 93.75% for Chinese). Principal component analysis and cluster analysis using all of the assayed elements and compounds were also conducted to determine which elements significantly accounted for the variation of the taro mineral composition. We report on the potential of H(2)PO(4)(-) and Co concentrations to differentiate taro grown in China and Japan and discuss the sources of variability in the taro mineral composition of our samples.     

Objective evaluation of whiteness of cooked rice and rice cakes using a portable spectrophotometer

The whiteness of cooked rice and rice cakes was evaluated using a portable spectrophotometer with a whiteness index (WI). Also, by using boiled rice for measurement of Mido values by Mido Meter, it was possible to infer the whiteness of cooked rice without rice cooking. In the analysis of varietal differences of cooked rice, ‘Tsuyahime’, ‘Koshihikari’ and ‘Koshinokaori’ showed high whiteness, while ‘Satonoyuki’ had inferior whiteness. The whiteness of rice cakes made from ‘Koyukimochi’ and ‘Dewanomochi’ was higher than the whiteness of those made from ‘Himenomochi’ and ‘Koganemochi’. While there was a significant correlation (r = 0.84) between WI values and whiteness scores of cooked rice by the sensory test, no correlation was detected between the whiteness scores and Mido values, indicating that the values obtained by a spectrophotometer differ from those obtained by a Mido Meter. Thus, a spectrophotometer may be a novel device for measurement of rice eating quality.     

Involvement of endogenous vasopressin in high plasma osmolality-induced anorexia via V1 receptor-mediated mechanism

It is known that water deprivation or injection of hypertonic saline induces anorexia. The present study examined the possible involvement of vasopressin in the suppression of food intake during high plasma osmolality. Intraperitoneal injection of vasopressin (20 microg/kg) into male rats significantly suppressed food intake for 1 hr. This anorectic effect of vasopressin was reversed by simultaneous injection of a peptide antagonist for V(1) receptor (40 microg/kg), but not for V(2) receptor (40 microg/kg). Intraperitoneal injection of hypertonic saline (20% NaCl, 2 ml/kg) similarly suppressed food intake for 2 hr, which was associated with a transient increase in plasma vasopressin concentrations. This hypertonic saline-induced suppression of food intake was blocked by a V(1) receptor antagonist. Vasopressin (40 ng/2 microl) directly administered into the third ventricle of the brain also suppressed food intake for 1 hr. These results suggest that vasopressin participates in the suppression of food intake during high plasma osmolality, the action of which is mediated by V(1) receptors in the brain.     

Loss of p16/Ink4a drives high frequency of rhabdomyosarcoma in a rat model of Duchenne muscular dystrophy

Rhabdomyosarcoma (RMS) is an aggressive type of soft tissue sarcoma, and pleomorphic RMS is a rare subtype of RMS found in adult. p16 is a tumor suppressor which inhibits cell cycle. In human RMS, p16 gene is frequently deleted, but p16-null mice do not develop RMS. We reported that genetic ablation of p16 by the crossbreeding of p16 knock-out rats (p16-KO rats) improved the dystrophic phenotype of a rat model of Duchenne muscular dystrophy (Dmd-KO rats). However, p16/Dmd double knock-out rats (dKO rats) unexpectedly developed sarcoma. In the present study, we raised p16-KO, Dmd-KO, and dKO rats until 11 months of age. Twelve out of 22 dKO rats developed pleomorphic RMS after 9 months of age, while none of p16-KO rats and Dmd-KO rats developed tumor. The neoplasms were connected to skeletal muscle tissue with indistinct borders and characterized by diffuse proliferation of pleomorphic cells which had eosinophilic cytoplasm and atypical nuclei with anisokaryosis. For almost all cases, the tumor cells immunohistochemically expressed myogenic markers including desmin, MyoD, and myogenin. The single cell cloning from tumor primary cells gained 20 individual Pax7-negative MyoD-positive RMS cell clones. Our results demonstrated that double knock-out of p16 and dystrophin in rats leads to the development of pleomorphic RMS, providing an animal model that may be useful to study the developmental mechanism of pleomorphic RMS.     

Analysis of Potassium Uptake by Rice Roots Treated with Aluminum Using a Positron Emitting Nuclide, 38K

We investigate the effect of Al on K⁺ uptake by rice roots. Potassium-38 (³⁸K), a positron emitting nuclide (the half-life: 7.61 min), was used to trace K⁺ behavior. When a rice root was treated with 10μM Al for 24 h, the uptake of ³⁸K in the root was increased in the range of 1 to 2 cm from the root tip compared with that of the control sample. Because the root continued to grow without showing any damage of plasma membrane during the Al treatment, it was suggested that the ³⁸K uptake was not occurred through diffusion into the cells. The uptake of ³⁸K in both treatments, with/without Al, was decreased by VO₄^3- (inhibitor of H⁺-ATPase on plasma membrane) and DNP (H⁺ ionophore) treatment, which suggested that the K⁺ uptake was performed through an active transport, such as H⁺:K⁺ transport or H⁺ gradient promoted by an Al treatment.