Analysis of Electrocardiograms and Behavior in Mice from Pregnancy to Lactation Period.

Changes in the mother-offspring relationship are presumably accompanied by dynamic changes in the autonomic nervous system. Although temporal measurements of autonomic activity have been performed in human mothers and infants, the analysis of long-term changes remains unexplored. Mouse mothers can form social bonds with their pups and have a short period of pregnancy and lactation, which makes them useful for the examination of physiological changes from pregnancy to pup-rearing. Therefore, a telemetry system was used for several weeks to measure the changes in the autonomic nervous system and the behavior of mouse mothers. The current results showed that an electrocardiogram (ECG) could be stably recorded regardless of the movements of mothers and parturition. ECG analysis showed that the heart rate gradually decreased from pregnancy to lactation, and sympathetic activity sharply increased as the pups developed. Furthermore, the simultaneous recording of behavior and ECG in the home cage enabled us to understand the behavior-dependent influences on the ECG, thereby revealing the characteristics of autonomic nervous activity during each behavior. Thus, the present experimental method helps to understand how the physiological characteristics of mothers change from pregnancy through pup rearing, supporting the healthy development of pups.

DNMT1/PKR double knockdowned HepG2 (HepG2-DP) cells have high hepatic function and differentiation ability.

HepG2 cells are widely used as a human hepatocytes model, but their functions, including drug metabolism, are inferior to primary hepatocytes. We previously reported that the hepatic gene expressions in HepG2 cells were upregulated by treatment with zebularine, which is an inhibitor of DNA methylation, through the inhibition of both DNA methyltransferase 1 (DNMT1) and double-stranded RNA-dependent protein kinase (PKR). In this study, we established a new HepG2 cell subline, HepG2-DP cells, by stable double knockdown of DNMT1 and PKR and evaluated its function. Albumin production, expression of CYP1A2 genes, and accumulation of lipid droplets were increased in HepG2-DP cells compared with the original HepG2 cells. Comprehensive gene expression analysis of transcription factors revealed that the expression of important genes for hepatic function, such as HNF1ß, HNF4α, ONECUT1, FOXA1, FOXA2, FOXA3, and various nuclear receptors, was upregulated in HepG2-DP cells. These results indicate that the newly established HepG2-DP cells are a highly functional hepatocyte cell line. In addition, we investigated whether HepG2-DP cells are able to mature by differentiation induction, since HepG2 cells are derived from hepatoblastoma. The gene expression of major CYPs and Phase II, III drug-metabolizing enzyme genes was significantly increased in HepG2-DP cells cultured in differentiation induction medium. These results suggest that HepG2-DP cells can be further matured by the induction of differentiation and could therefore be applied to studies of drug metabolism and pharmacokinetics.

Difference in infantile mother preference is associated with adult anxiety/object recognition in C57BL/6 mice.

We previously reported that mice pups showed individual differences in mother preferences at 16 days old; some pups show high preference to their mother, and other pups show low preference to it. In this study, we examined whether these individual differences were associated with anxiety-like behavior and cognition functions in adulthood. We found that pups showing low mother preference exhibit low anxiety-like behavior and impaired object cognition in adulthood. These results suggest that some behavioral characteristics in adulthood may be associated with the profile of mother preference prior to weaning.

Low-density cell culture enhances hepatic function through tight junction formation in HepG2 cells.

BACKGROUND INFORMATION: An in vitro evaluation system using cultured hepatocytes is the most useful method in preclinical research, such as drug metabolism and toxicity test. Human hepatocytes should be used in an in vitro evaluation system because the expression of drug-metabolizing enzymes varies among animal species. HepG2 cells, a liver cancer-derived cell line, are widely used as a human hepatocyte model; however, their hepatic functions are generally weak. RESULTS: In this study, we showed that low-density HepG2 cell culture induces hepatic function. The morphology of HepG2 cells was altered depending on the cell density at the time of seeding. Low-density cultured HepG2 cells proliferated as tightly packed colonies. The HepG2 cell colonies in low-density culture demonstrated enhanced tight junction formation. Tight junction protein gene expression levels, such as those of zonula occludens-1 (ZO-1), junctional adhesion molecule 1 (JAM), claudin, occludin, and tricellulin, increased in low-density cultured HepG2 cells. Phases I and II metabolic enzymes, phase III transporter gene expression, and CYP3A4 activity also increased in low-density cultured HepG2 cells. Occludin and tricellulin knockdown inhibited the increased hepatic function in low-density cultures. Tricellulin knockdown reduced the expression of hepatocyte nuclear factor 6 (HNF6), CCAAT/enhancer-binding protein alpha (CEBPA), and aryl hydrocarbon receptor (AHR). In addition, the expression of nuclear receptor subfamily 1 group h member 2 (NR1H2) increased in low-density cultures, canceled by occludin and tricellulin knockdown. CONCLUSIONS: Our results suggest that low-density HepG2 cell cultures enhance hepatic function by promoting tight junction formation and demonstrate the importance of cell density in drug evaluation using hepatocyte cell lines.

Relationship between infantile mother preference and neural regions activated by maternal contact in C57BL/6 mice.

Mutual attachment between mother and pup is important to enable the mother to care for her pup and for the pup to receive care from its mother. Pups eventually leave their mothers, which is also very important to their growth. The mechanism of preference by which pups transfer attachment from their mother to others remains unknown. In this study, we assessed mother/novel dam preferences and examined the brain regions associated with the regulation of this preference in C57BL/6 mice pups. We found that C57BL/6 mice pups had variety in their mother/novel dam preferences at 16 days old. This variety was not related to the sex of the pups, their weight, or the litter size. In order to clarify the brain mechanisms responsible for this variety, we examined the relationship between mother/novel dam preference and neuronal activation induced by contact with the mother. We found that pups exhibiting novel dam preference had higher neural activity in the anterior cingulate cortex (ACC) and bed nucleus of the stria terminalis (BNST) when exposed to their mother. These results suggest that ACC and/or BNST neural activity may be associated with mother/novel dam preferences in infant mice.

Associations among amino acid, lipid, and glucose metabolic profiles in childhood obesity.

BACKGROUND: Plasma-free amino acid profiles have been reported to correlate with obesity and glucose metabolism, and have been studied as potentially useful biomarkers of lifestyle-related diseases affecting metabolism in adulthood. However, knowledge of these relationships is lacking in children, despite the growing public health problem posed by childhood obesity. The aim of this study was to assess whether plasma-free amino acid profiles can serve as useful biomarkers of lifestyle-related diseases in children with obesity. METHODS: This retrospective study used the medical records of 26 patients (15 male, 11 female) aged 9 or 10 years presenting with moderate to severe obesity and hyperlipidemia between April 2015 and March 2017. A degree of obesity of 30% or more was defined as moderate or severe. Amino acid levels were compared between obese children with and without impaired glucose tolerance using a t-test or Mann-Whitney U test. In addition, the influence of factors such as intima media thickness, low-density lipoprotein cholesterol, high-density lipoprotein cholesterol, amino acids, and homeostasis model assessment-insulin resistance (HOMA-IR) were analyzed pairwise using Pearson's correlation or Spearman's rank correlation. RESULTS: HOMA-IR was positively correlated with valine, leucine (Leu), isoleucine, phenylalanine, tryptophan, methionine, threonine, lysine, alanine, tyrosine, glutamate (Glu), proline, arginine, ornithine, total free amino acids (all P < 0.01), and aspartate (P = 0.010). Moreover, blood uric acid levels were positively correlated with Leu (P = 0.005) and Glu (P = 0.019), and negatively correlated with serine, glycine, and asparagine (P = 0.007, P = 0.003, and P = 0.013, respectively). CONCLUSIONS: Amino acid profile reflects impaired glucose tolerance and hyperuricemia at an early stage of obesity. It is therefore a useful marker to inform early intervention in children with obesity, as in adults.

Modulation of male mouse sociosexual and anxiety-like behaviors by vasopressin receptors.

Although the involvement of two types of vasopressin (AVP) receptors, v1a and v1b, in neural regulation of social behavior is well documented in rodents, there is no report on combined actions of them in regulation of social behavior. In this study, we investigated behavioral differences between wild-type (WT) and v1a and v1b double knockout (dKO) mice. For this, we measured olfactory preference, sexual behavior with receptive females (four weekly tests) in an enriched large observation cage, and anxiety-like behaviors. No difference between WT and dKO mice was found in olfactory preferences for estrous female odor to male odor. Over all four mating tests, the number of mounts and pursuits after receptive females was significantly greater in dKO mice than in WT mice. In the elevated plus maze and the open field test, dKO mice showed lower anxiety-like behavior than WT mice. Finally, we measured approach behavior to several types of objects, figurines, and caged anestrous or estrous females placed in the open field apparatus. The only difference observed was that dKO mice spent longer in the vicinity of estrous females than did WT mice. These findings suggest that vasopressin receptors are involved in the regulation of sociosexual behavior, presumably partly mediated by emotional responses, in male mice.

Reducing artifacts of gadoxetate disodium-enhanced MRI with oxygen inhalation in patients with prior episode of arterial phase motion: intra-individual comparison.

PURPOSE: To determine whether oxygen inhalation reducing artifacts in patients with previous transient severe motion (TSM) on gadoxetate-disodium-enhanced MRI. MATERIALS AND METHODS: Fifty-one patients with TSM on previous gadoxetate-disodium-enhanced MRI (Baseline examination) were evaluated. Image quality in the examination with oxygen inhalation (Oxygen examination) and that in Baseline examination and the examination before Baseline examination without oxygen inhalation (Past examination) were qualitatively compared in dynamic study. RESULTS: Image quality was significantly higher in Oxygen examination than Baseline examination in arterial phase, but there was no statistical difference between Baseline and Past examinations. CONCLUSION: Oxygen inhalation improved image quality in patients with a prior episode of arterial phase TSM.

Measuring hepatic functional reserve using T1 mapping of Gd-EOB-DTPA enhanced 3T MR imaging: A preliminary study comparing with 99mTc GSA scintigraphy and signal intensity based parameters.

PURPOSE: To determine the utility of liver T1-mapping on gadolinium-ethoxybenzyl-diethylenetriamine-pentaacetic acid (Gd-EOB-DTPA) enhanced magnetic resonance (MR) imaging for the measurement of liver functional reserve compared with the signal intensity (SI) based parameters, technetium-99m-galactosyl serum albumin (99mTc-GSA) scintigraphy and indocyanine green (ICG) clearance. MATERIALS AND METHODS: This retrospective study included 111 patients (Child-Pugh-A 90; -B 21) performed with both Gd-EOB-DTPA enhanced liver MR imaging and 99mTc-GSA (76 patients with ICG). Receiver operating characteristic (ROC) curve analysis was performed to compare diagnostic performances of T1-relaxation-time parameters [pre-(T1pre) and post-contrast (T1hb) Gd-EOB-DTPA], SI based parameters [relative enhancement (RE), liver-to-muscle-ratio (LMR), liver-to-spleen-ratio (LSR)] and 99mTc-GSA scintigraphy blood clearance index (HH15)] for Child-Pugh classification. Pearson's correlation was used for comparisons among T1-relaxation-time parameters, SI-based parameters, HH15 and ICG. RESULTS: A significant difference was obtained for Child-Pugh classification with T1hb, ΔT1, all SI based parameters and HH15. T1hb had the highest AUC followed by RE, LMR, LSR, ΔT1, HH15 and T1pre. The correlation coefficients with HH15 were T1pre 0.22, T1hb 0.53, ΔT1 -0.38 of T1 relaxation parameters; RE -0.44, LMR -0.45, LSR -0.43 of SI-based parameters. T1hb was highest for correlation with HH15. The correlation coefficients with ICG were T1pre 0.29, T1hb 0.64, ΔT1 -0.42 of T1 relaxation parameters; RE -0.50, LMR -0.61, LSR -0.58 of SI-based parameters; 0.64 of HH15. Both T1hb and HH15 were highest for correlation with ICG. CONCLUSION: T1 relaxation time at post-contrast of Gd-EOB-DTPA (T1hb) was strongly correlated with ICG clearance and moderately correlated HH15 with 99mTc-GSA. T1hb has the potential to provide robust parameter of liver functional reserve.

Hepatic fat quantification using automated six-point Dixon: Comparison with conventional chemical shift based sequences and computed tomography.

PURPOSE: To compare automated six-point Dixon (6-p-Dixon) MRI comparing with dual-echo chemical-shift-imaging (CSI) and CT for hepatic fat fraction in phantoms and clinical study. MATERIALS AND METHODS: Phantoms and fifty-nine patients were examined both MRI and CT for quantitative fat measurements. RESULTS: In phantom study, linear regression between fat concentration and 6-p-Dixon showed good agreement. In clinical study, linear regression between 6-p-Dixon and dual-echo CSI showed good agreement. CT attenuation value was strongly correlated with 6-p-Dixon (R2=0.852; P<0.001) and dual-echo CSI (R2=0.812; P<0.001). CONCLUSION: Automated 6-p-Dixon and dual-echo CSI were accurate correlation with CT attenuation value of liver parenchyma. 6-p-Dixon has the potential for automated hepatic fat quantification.

Uniform vascular enhancement of lower-extremity artery on CT angiography using test-injection monitoring at the central level of the scan range: a simulation flow phantom study with clinical correlation.

RATIONALE AND OBJECTIVES: To evaluate the efficacy of variable contrast injection durations and scanning delay determined by test injection analysis of computed tomography angiography (CTA) of peripheral arteries. MATERIALS AND METHODS: We used a flow phantom that simulates the hemodynamics in a lower extremity artery. We set the flow rate at the pump to 2.0 or 5.0 L/minute. In protocol 1, we adopted a variable contrast injection duration based on the peak enhancement time of the test injection monitoring at the central level of the scan range. In protocol 2, we adopted a fixed contrast injection duration. The scanning delay was determined with a conventional bolus-tracking technique monitoring at the top of the scan range. Mean arterial attenuation and difference between the maximum and minimum attenuation values were calculated. To verify the phantom study results, clinical study, including 16 patients was performed under protocol 1. RESULTS: The mean attenuation values under protocols 1 and 2 were comparable (563.6 Hounsfield units [HU] and 535.0 HU, respectively) at a pump flow rate of 2.0 L/minute; at 5.0 L/minute, they were 289.4 HU and 328.8 HU. The difference between the maximum and minimum attenuation values was smaller under protocol 1 than protocol 2 (76.8 HU vs. 184.9 HU) at a pump flow of 2.0 L/minute and also smaller under protocol 1 than protocol 2 (79.7 HU vs. 203.8 HU) at 5.0 L/minute. In clinical study, the mean attenuation value was 332.6 +/- 51.9 HU, and the difference between the maximum and minimum attenuation values was 55.1 +/- 24.4 HU. CONCLUSION: The object-specific injection duration based on test injection at the central level of the scan range provides sufficient and constant vascular enhancement at CTA.