Regulation of ghrelin secretion during pregnancy and lactation in the rat: possible involvement of hypothalamus

We investigated the plasma concentration of ghrelin peptide during pregnancy and lactation in rats. Plasma ghrelin levels on days 10 and 15 of pregnancy were significantly lower than those of the non-pregnant rats. Thereafter, the plasma ghrelin levels on day 20 of pregnancy sharply increased to levels comparable with those in non-pregnant rats. Ghrelin peptide concentrations in the stomach did not change significantly during pregnancy. In the hypothalamus, ghrelin mRNA levels were significantly lower on day 15 of pregnancy than in the non-pregnant rats. Also, plasma ghrelin levels were significantly lower in lactating dams than non-lactating controls on days 3 and 8 of lactation. We examined the possible involvement of prolactin and oxytocin in the regulation of plasma ghrelin concentrations during lactation. Although plasma prolactin levels were decreased by the administration of bromocriptine, plasma ghrelin levels did not differ significantly between vehicle- and drug-treated lactating rats. Administration of haloperidol produced a marked increase in plasma prolactin levels as compared with the non-lactating controls. However, plasma ghrelin levels were not significantly different between vehicle- and drug-treated rats. Administration of an oxytocin antagonist into the lateral ventricle significantly inhibited the increase in the plasma oxytocin level induced by acute suckling. However, plasma ghrelin levels did not significantly between the groups. These observations indicated that the decrease in serum ghrelin is caused by a loss of the contribution of hypothalamic ghrelin. Furthermore, the present results suggested that the suckling stimulus itself, but the release of prolactin or oxytocin, is the factor most likely to be responsible for the suppression of ghrelin secretion during lactation.     

2-Ureidoquinoline: a useful molecular element for stabilizing single cytosine and thymine bulges

We have demonstrated that aromatic heterocycles having hydrogen-bonding surfaces complementary to those of nucleotide bases are effective molecular elements for the binding to single nucleotide bulges and base mismatches. We here report that a new molecule, 2-ureidoquinoline having an alignment of hydrogen-bonding groups in the order of acceptor-donor-donor stabilizes single cytosine and thymine bulges in duplex DNAs. Furthermore, a dimeric form of 2-ureidoquinoline stabilizes cytosine-cytosine and cytosine-thymine mismatches.     

Secular trends of sizes at birth in Japanese healthy infants born between 1962 and 1988

Body sizes at birth are important clinical indicators widely used for evaluation of prenatal growth. Japan had significant socioeconomic improvement around the 1960s, and these environmental changes may influence physiologically prenatal growth. Furthermore, in Japan, measurements of size at birth for birth certificates are weight and height. Thus, we can refer to annual data on weight and height, but not on head and chest circumference at birth. In this study we measured the weight, height, and head and chest circumference at birth among 6,563 Japanese singleton healthy infants, annually in 1962 and 1988, and examined secular trends of these anthropometric measurements. The boys consistently exceeded the girls in all four variables. Birth weight and height increased significantly from the 1960s to '70s, but did not differ between the '70s and '80s in both boys and girls. Secular trends of head and chest circumference were different from them. In both boys and girls, head and chest circumference increased significantly from the '60s to the '70s, but decreased significantly from the '70s to the '80s. No difference of head circumference during the '60s and '80s was found, but the difference of chest circumference was found. Size at birth was likely to increase from the '60s to '70s in Japan. These findings suggest that the environmental changes such as socioeconomic improvements influence the prenatal growth.     

Cell-passage activity is required for the malarial parasite to cross the liver sinusoidal cell layer

Liver infection is an obligatory step in malarial transmission, but it remains unclear how the sporozoites gain access to the hepatocytes, which are separated from the circulatory system by the liver sinusoidal cell layer. We found that a novel microneme protein, named sporozoite microneme protein essential for cell traversal (SPECT), is produced by the liver-infective sporozoite of the rodent malaria parasite, Plasmodium berghei. Targeted disruption of the spect gene greatly reduced sporozoite infectivity to the liver. In vitro cell invasion assays revealed that these disruptants can infect hepatocytes normally but completely lack their cell passage ability. Their apparent liver infectivity was, however, restored by depletion of Kupffer cells, hepatic macrophages included in the sinusoidal cell layer. These results show that malarial sporozoites access hepatocytes through the liver sinusoidal cell layer by cell traversal motility mediated by SPECT and strongly suggest that Kupffer cells are main routes for this passage. Our findings may open the way for novel malaria transmission-blocking strategies that target molecules involved in sporozoite migration to the hepatocyte.     

Novel biodegradable polyphosphate cross-linker for making biocompatible hydrogel

To obtain a novel biodegradable cross-linker, polymerizable polyphosphate (PIOP) was synthesized by ring-opening polymerization of 2-i-propyl-2-oxo-1,3,2-dioxaphospholane with 2-(2-oxo-1,3,2-dioxaphosphoroyloxyethyl methacrylate) (OPEMA). The number averaged molecular weight of the PIOP was 1.2 x 10(4), and the number of OPEMA units in one PIOP molecule was 2.2. Nonenzymatic degradation of the PIOP was evaluated in various pH aqueous media. The degree of hydrolysis was dependent on the pH; that is, it increased with an increase in the pH of the medium. At pH 11.0, the PIOP completely degraded in only 6 days. The poly[2-methacryloyloxyethyl phosphorylcholine (MPC)] cross-linked with the PIOP was prepared by radical polymerization. This polymer could form hydrogel, and the free water fraction in the hydrogel was high. The enzymatic activity of trypsin in contact with the hydrogel was similar to that in buffer solution. There is no adverse effect caused by the hydrogel to reduce the function of the trypsin. The cytotoxicity of poly(MPC) and degraded PIOP was evaluated using v79 cells, and it was not observed in either case. In conclusion, PIOP is a hydrolyzable polymer, which can be used as a cross-linker, and novel hydrogels having biodegradability and biocompatibility were prepared from poly(MPC) cross-linked with the PIOP.     

Short-term ascorbic acid deficiency induced oxidative stress in the retinas of young guinea pigs

We examined whether short-term ascorbic acid deficiency induces oxidative stress in the retinas of young guinea pigs. Four-week-old guinea pigs were given a scorbutic diet (20 g/animal/day) with and without adequate ascorbic acid (400 mg/animal/day) in drinking water for 3 weeks. The serum concentrations of the reduced form of ascorbic acid and the oxidized form of ascorbic acid in the deficient group were 14.1 and 4.1%, respectively, of those in the adequate group. The retinal contents of the reduced form of ascorbic acid and the oxidized form of ascorbic acid in the deficient group were 6.4 and 27.3%, respectively, of those in the adequate group. The retinal content of thiobarbituric acid-reactive substances, an index of lipid peroxidation, was 1.9-fold higher in the deficient group than in the adequate group. Retinal reduced glutathione and vitamin E contents in the deficient group were 70.1 and 69.4%, respectively, of those in the adequate group. This ascorbic acid deficiency did not affect serum thiobarbituric acid-reactive substances and reduced glutathione concentrations but increased serum vitamin E concentration. These results indicate that short-term ascorbic acid deficiency induces oxidative stress in the retinas of young guinea pigs without disrupting systemic antioxidant status.     

Uniaxial strain upregulates matrix-degrading enzymes produced by human vascular smooth muscle cells

Arteries remodel in response to environmental changes. We investigated whether mechanical strain modulates production of matrix metalloproteinase (MMP)-2 and -9 by cultured vascular smooth muscle cells (SMC). MMP-2 and MMP-9 expression were tested using human saphenous vein SMC cultured on silicone membranes at rest or subjected to physiological levels (5%) of stationary or cyclical (1 Hz) uniaxial strain. Compared with control, stationary strain significantly increased MMP-2 mRNA levels at all time points, whereas cyclic strain decreased it after 48 h. Both secreted and cell-associated pro-MMP-2 levels were increased by stationary strain at all times (P < 0.01), whereas cyclic strain decreased secreted levels after 48 h (P < 0.02). MMP-9 mRNA levels and pro-MMP-9 protein were increased after 48 h of stationary stretch (P < 0.01) compared with both no strain and cyclic strain. Our study indicates that vascular SMC show a selective response to different types of strain. We suggest that local increases in stationary mechanical strain resulting from stenting, hypertension, or atherosclerosis may lead to enhanced matrix degradation by SMC.     

Production of singlet oxygen on irradiation of a photodynamic therapy agent, zinc-coproporphyrin III, with low host toxicity

Zinc-coproporphyrin III (Zincphyrin) acts efficiently as a photodynamic therapy (PDT) agent in mice, while it shows no tumor cell-killing activity in vitro and has a high LD₅₀ (low toxicity) in mice. It appears to have advantages over other porphyrins as a practical PDT reagent. In order to examine the action mechanism of Zincphyrin in PDT, we evaluated the photochemical characteristics of Zincphyrin by measurement of the near-infrared emission at 1268 nm, which provides direct evidence for formation of ¹O₂. Intense emission was observed in the presence of Zincphyrin, and was completely inhibited by NaN₃, a ¹O₂ scavenger. Based on a quenching study, the rate constant of the reaction of ¹O₂ with NaN₃ was determined to be 1.5–3.5 M^–1 s^–1, which is close to the reported value (3.8×10⁸ M^–1 s^–1). The intensity of the ¹O₂-specific emission was proportional to both the laser power and the concentration of Zincphyrin. The fluorescence quantum yield of Zincphyrin was 0.004 in phosphate buffer (100 mM, pH 7.4), which indicates that the excited state decays via other pathway(s) faster than through the fluorescence emission pathway. The lifetime of the triplet state of Zincphyrin (210 s) was relatively long compared to that of other porphyrins, such as hematoporphyrin (Hp) (40 s), coproporphyrin I (50 s), or coproporphyrin III (36 s). These results demonstrate the photodynamic generation of ¹O₂ by Zincphyrin.     

Proteasome activator PA28gamma-dependent nuclear retention and degradation of hepatitis C virus core protein

Hepatitis C virus (HCV) core protein plays an important role in the formation of the viral nucleocapsid and a regulatory protein involved in hepatocarcinogenesis. In this study, we have identified proteasome activator PA28gamma (11S regulator gamma) as an HCV core binding protein by using yeast two-hybrid system. This interaction was demonstrated not only in cell culture but also in the livers of HCV core transgenic mice. These findings are extended to human HCV infection by the observation of this interaction in liver specimens from a patient with chronic HCV infection. Neither the interaction of HCV core protein with other PA28 subtypes nor that of PA28gamma with other Flavivirus core proteins was detected. Deletion of the PA28gamma-binding region from the HCV core protein or knockout of the PA28gamma gene led to the export of the HCV core protein from the nucleus to the cytoplasm. Overexpression of PA28gamma enhanced the proteolysis of the HCV core protein. Thus, the nuclear retention and stability of the HCV core protein is regulated via a PA28gamma-dependent pathway through which HCV pathogenesis may be exerted.