Effects of amino acids and albumin administration on albumin metabolism in surgically stressed rats: A basic nutritional study.

BACKGROUND: Nutrition therapy and administration of albumin preparations are common in postsurgical patients. However, the effects of these interventions on albumin metabolism are unclear. We elucidated the effect of postoperative albumin and/or parenteral nutrition administration on it. METHODS: Sprague-Dawley rats underwent surgery involving intestinal rubbing followed by intestinal exposure. Subsequently, they were administered experimental solutions for 48 h, their blood samples were collected at 24 and 48 h, and livers were excised at 48 h. Based on experimental solutions, rats were divided into five groups: non-surgical (Non-surg); glucose and electrolyte solution (GE); amino acid, glucose, and electrolyte solution (AGE); GE + rat serum albumin (Alb) (GE + Alb); and AGE + Alb. Their plasma albumin concentrations; albumin fractional synthesis rate (ALB FSR); mercaptoalbumin/total albumin ratio (MA ratio); and messenger RNA (mRNA) expressions of albumin and hepatocyte nuclear factor-1 (HNF-1) in the liver were measured. RESULTS: The GE and AGE groups showed significant decline in albumin concentrations. ALB FSR was significantly enhanced in the AGE group compared with the GE group. The mRNA expression of albumin was similar to ALB FSR in all groups and that of HNF-1 was significantly decreased in the GE + Alb and AGE + Alb groups compared with the Non-surg group. The MA ratio in the AGE group was similar to the Non-surg group. CONCLUSION: The administration of amino acids comprising parenteral nutrition after surgery augmented ALB FSR and maintained the MA ratio only without simultaneous albumin administration.

Effects of neonatal rearing by different types of foster mother on the distribution of corticotropin-releasing factor neurons in the central amygdaloid nucleus in rats.

The mother-child relationship of newborns plays an essential role in the development of the central nervous system, and an inadequate relationship, such as mother-child separation, can cause deficits of mental function in adulthood. However, insufficient research has examined the effects of foster mothers. We assigned some neonatal rats to one of two foster mothers: one that was lactating and feeding her first litter (FL group) and one that had one previous experience of childbirth and feeding but no current litter (FE group). Other pups were raised by their own mother (OM group) or subjected to maternal separation (MS group). Pups were placed with the foster mother (FL and FE groups) or separated from their mother (MS group) for 3 h/day on postnatal days 1-20. At age 6 weeks, each group was divided into two subgroups, one with 30 min of acute restraint stress loading (FL-R, FE-R, OM-R, and MS-R) and one without it (FL, FE, OM, and MS). Then, we compared the density of corticotropin-releasing factor-immunoreactive (CRF-ir) neurons in the central amygdaloid nucleus (CeA). The density of CRF-ir neurons in the CeA was significantly lower in the FL-R and MS-R subgroups than in the FL and MS subgroups, respectively. The results suggest that differences in care received during the neonatal period affect maturation of CRF neurons in the CeA and may have negative effects on the synthesis and release of CRF.

Anti-inflammatory Effect of the Water-Soluble Portion of Porcine Placental Extract in Lipopolysaccharide-Stimulated RAW264.7 Murine Macrophage Cells.

Porcine placental extract (PPE) is used as a nonprescription drug for analeptics and in health foods and cosmetics in Japan, Korea and China. It was reported that PPE has anti-oxidative and anti-inflammatory activities; however, the mechanisms and the responsible molecules involved in these activities are still unclear. Here, we investigated how enzymatically prepared PPE affects proinflammatory factors such as interleukin (IL)-1ß, IL-6, and tumor necrosis factor (TNF)-α in a cultured macrophage cell line, RAW264.7, when co-stimulated with lipopolysaccharide (LPS). Enhanced production of IL-1ß, IL-6 and TNF-α by LPS was significantly reduced by the addition of PPE and these effects were dose dependent. Nitric oxide (NO) production induced in cultured macrophages by LPS was also inhibited by PPE. Real-time PCR after the reverse transcription of total RNAs isolated from cells treated with PPE revealed that the mRNA expressions of IL-1ß, IL-6, TNFα, and NO synthase (NOS)-2 were reduced. The necessary concentration of PPE prepared by enzymatic digestion to mediate anti-inflammatory effects compared with the reported value of that extracted by phosphate buffered saline without digestion was proportional to the amount of extracted materials from the same amount of placenta (about 10-fold). This suggests that the molecules responsible for the anti-inflammatory activity exists in the placenta and can be extracted by phosphate buffered saline, and thus might survive enzymatic digestion.

A spinach O-acetylserine(thiol)lyase homologue, SoCSaseLP, suppresses cysteine biosynthesis catalysed by other enzyme isoforms.

An enzyme, O-acetylserine(thiol)lyase (OASTL), also known as O-acetylserine sulfhydrylase or cysteine synthase (CSase), catalyses the incorporation of sulfide into O-acetylserine and produces cysteine. We previously identified a cDNA encoding an OASTL-like protein from Spinacia oleracea, (SoCSaseLP), but a recombinant SoCSaseLP produced in Escherichia coli did not show OASTL activity. The exon-intron structure of the SoCSaseLP gene shared conserved structures with other spinach OASTL genes. The SoCSaseLP and a Beta vulgaris homologue protein, KMT13462, comprise a unique clade in the phylogenetic tree of the OASTL family. Interestingly, when the SoCSaseLP gene was expressed in tobacco plants, total OASTL activity in tobacco leaves was reduced. This reduction in total OASTL activity was most likely caused by interference by SoCSaseLP with cytosolic OASTL. To investigate the possible interaction of SoCSaseLP with a spinach cytosolic OASTL isoform SoCSaseA, a pull-down assay was carried out. The recombinant glutathione S-transferase (GST)-SoCSaseLP fusion protein was expressed in E. coli together with the histidine-tagged SoCSaseA protein, and the protein extract was subjected to glutathione affinity chromatography. The histidine-tagged SoCSaseA was co-purified with the GST-SoCSaseLP fusion protein, indicating the binding of SoCSaseLP to SoCSaseA. Consistent with this interaction, the OASTL activity of the co-purified SoCSaseA was reduced compared with the activity of SoCSaseA that was purified on its own. These results strongly suggest that SoCSaseLP negatively regulates the activity of other cytosolic OASTL family members by direct interaction.

Anucleate cell blue assay: a useful tool for identifying novel type II topoisomerase inhibitors.

About 95,000 compounds were screened by the anucleate cell blue assay. Fifty-one of the hit compounds had various structures and showed inhibitory activity against DNA gyrase and/or topoisomerase IV. Moreover, the compounds exhibited antibacterial activity against a fluoroquinolone- and novobiocin-resistant strain of Staphylococcus aureus. The anucleate cell blue assay is therefore a useful tool for finding novel type II topoisomerase inhibitors.

Accumulation of mutations in both gyrB and parE genes is associated with high-level resistance to novobiocin in Staphylococcus aureus.

Coumarin-resistant mutants of Staphylococcus aureus were isolated by three-step selection with novobiocin at different concentrations. Sequencing analysis of the gyrB and parE genes of the first-, second-, and third-step mutants revealed that successive point mutations first occurred specifically in the gyrB gene, followed by a point mutation in the parE gene and then an additional point mutation in the gyrB gene. These findings demonstrate that DNA gyrase is the primary target and that topoisomerase IV is the secondary target for novobiocin and that the accumulation of point mutations in both the gyrB and the parE genes is associated with high-level resistance to novobiocin in S. aureus. Moreover, our results show that the amino acid substitutions (Asp-89 to Gly and Ser-128 to Leu) found in GyrB are associated with resistance to novobiocin but not to coumermycin A1, suggesting that the interactions of novobiocin and coumermycin A1 with GyrB differ at the molecular level.

Excitotoxic injury induces production of monocyte chemoattractant protein-1 in rat cortico-striatal slice cultures.

The effect of excitotoxic injury on the production of monocyte chemoattractant protein-1 (MCP-1) was examined in rat cortico-striatal slice cultures. Treatment with 50 microM N-methyl-D-aspartate (NMDA) for 4 h, which caused severe damage in neurons, induced the production of MCP-1 in astrocytes. Production levels were markedly elevated immediately after the treatment, peaked at 4-8 h, and had decreased to nearly the basal level by 72 h. Since the treatment promoted the release of MCP-1 in the slice cultures, but not in the enriched astrocyte cultures, it is unlikely that NMDA directly acted on the astrocytes. These results suggest that information on neuronal injury induced by NMDA is transmitted to astrocytes to induce the production of MCP-1. Organotypic slice cultures are useful for investigating the inflammatory responses of astrocytes in the process of neuronal injury.