Desynchronization between Food Intake and Light Stimulations Induces Uterine Clock Quiescence in Female Mice.

BACKGROUND: Dysmenorrhea is associated with breakfast skipping in young women, suggesting that fasting in the early active phase disrupts uterine functions. OBJECTIVES: To investigate the possible involvement of the uterine clock system in fasting-induced uterine dysfunction, we examined core clock gene expressions in the uterus using a 28-h interval-fed mouse model. METHODS: Young female mice (8 wk of age) were divided into 3 groups: group I (ad libitum feeding), group II (time-restricted feeding, initial 4 h of the active period every day), and group III (time-restricted feeding for 8 h with a 28-h cycle). Groups II and III have the same fasting interval of 20 h. After analyzing feeding and wheel running behaviors during 2 wk of dietary restriction, mice were sacrificed at 4-h intervals, and the expression profiles of clock genes in the uterus and liver were examined by qPCR. RESULTS: The mice in group I took food mainly during the dark phase and those in group II during the initial 4 h of the dark phase, whereas those in group III delayed feeding time by 4 h per cycle. In all groups, spontaneous wheel running was observed during the dark phase. There was no difference in the quantity of feeding and the amount of running exercise among the 3 groups during the second week. The mRNA expressions of peripheral clock genes, Bmal1, Clock, Per1, Per2, Cry1, Nr1d1, and Dbp and a clock-controlled gene, Fabp1, in the uterus showed rhythmic oscillations with normal sequential expression cascade in groups I and II, whereas their expressions decreased and circadian cycles disappeared in group III. In contrast, liver core clock genes in group III showed clear circadian cycles. CONCLUSIONS: Fluctuations in the timing of the first food intake impair the uterine clock oscillator system to reduce clock gene expressions and abolish their circadian rhythms.

The Circadian Clock, Nutritional Signals and Reproduction: A Close Relationship.

The circadian rhythm, which is necessary for reproduction, is controlled by clock genes. In the mouse uterus, the oscillation of the circadian clock gene has been observed. The transcription of the core clock gene period (Per) and cryptochrome (Cry) is activated by the heterodimer of the transcription factor circadian locomotor output cycles kaput (Clock) and brain and muscle Arnt-like protein-1 (Bmal1). By binding to E-box sequences in the promoters of Per1/2 and Cry1/2 genes, the CLOCK-BMAL1 heterodimer promotes the transcription of these genes. Per1/2 and Cry1/2 form a complex with the Clock/Bmal1 heterodimer and inactivate its transcriptional activities. Endometrial BMAL1 expression levels are lower in human recurrent-miscarriage sufferers. Additionally, it was shown that the presence of BMAL1-depleted decidual cells prevents trophoblast invasion, highlighting the importance of the endometrial clock throughout pregnancy. It is widely known that hormone synthesis is disturbed and sterility develops in Bmal1-deficient mice. Recently, we discovered that animals with uterus-specific Bmal1 loss also had poor placental development, and these mice also had intrauterine fetal death. Furthermore, it was shown that time-restricted feeding controlled the uterine clock's circadian rhythm. The uterine clock system may be a possibility for pregnancy complications, according to these results. We summarize the most recent research on the close connection between the circadian clock and reproduction in this review.

The regulation of glucose and lipid homeostasis via PLTP as a mediator of BAT-liver communication.

While brown adipose tissue (BAT) is well-recognized for its ability to dissipate energy in the form of heat, recent studies suggest multifaced roles of BAT in the regulation of glucose and lipid homeostasis beyond stimulating thermogenesis. One of the functions involves interorgan communication with metabolic organs, such as the liver, through BAT-derived secretory factors, a.k.a., batokine. However, the identity and the roles of such mediators remain insufficiently understood. Here, we employed proteomics and transcriptomics in human thermogenic adipocytes and identified previously unappreciated batokines, including phospholipid transfer protein (PLTP). We found that increased circulating levels of PLTP, via systemic or BAT-specific overexpression, significantly improve glucose tolerance and insulin sensitivity, increased energy expenditure, and decrease the circulating levels of cholesterol, phospholipids, and sphingolipids. Such changes were accompanied by increased bile acids in the circulation, which in turn enhances glucose uptake and thermogenesis in BAT. Our data suggest that PLTP is a batokine that contributes to the regulation of systemic glucose and lipid homeostasis as a mediator of BAT-liver interorgan communication.

Uterine Deletion of Bmal1 Impairs Placental Vascularization and Induces Intrauterine Fetal Death in Mice.

Recently, it was demonstrated that the expression of BMAL1 was decreased in the endometrium of women suffering from recurrent spontaneous abortion. To investigate the pathological roles of uterine clock genes during pregnancy, we produced conditional deletion of uterine Bmal1 (cKO) mice and found that cKO mice could receive embryo implantation but not sustain pregnancy. Gene ontology analysis of microarray suggested that uterine NK (uNK) cell function was suppressed in cKO mice. Histological examination revealed the poor formation of maternal vascular spaces in the placenta. In contrast to WT mice, uNK cells in the spongiotrophoblast layer, where maternal uNK cells are directly in contact with fetal trophoblast, hardly expressed an immunosuppressive NK marker, CD161, in cKO mice. By progesterone supplementation, pregnancy could be sustained until the end of pregnancy in some cKO mice. Although this treatment did not improve the structural abnormalities of the placenta, it recruited CD161-positive NK cells into the spongiotrophoblast layer in cKO mice. These findings indicate that the uterine clock system may be critical for pregnancy maintenance after embryo implantation.

Fish oil suppresses obesity more potently in lean mice than in diet-induced obese mice but ameliorates steatosis in such obese mice.

This study sought to clarify the antiobesity effects of fish oil (FO) in terms of prevention and amelioration. An isocaloric diet composed of lard or FO was given to lean C57BL/6J mice for the study of prevention and high-fat diet-induced obese (DIO) mice for the study of amelioration for 4 weeks. Body weight gain and food efficiency were potently suppressed by FO in lean mice compared to lard diet-fed mice. Uncoupling protein-1 (UCP-1) expression in inguinal white adipose tissue (WAT) was also significantly induced by FO in lean mice. FO also suppressed body weight gain and food efficiency in DIO mice but did not reduce body weight. FO ameliorated liver steatosis in DIO mice by mildly inducing UCP-1 in inguinal WAT. FO suppressed obesity more potently in lean mice than in DIO mice but ameliorated steatosis in the DIO mice.

The role of increased FGF21 in VLDL-TAG secretion and thermogenic gene expression in mice under protein malnutrition.

Protein malnutrition promotes hepatic lipid accumulation in growing animals. In these animals, fibroblast growth factor 21 (FGF21) rapidly increases in the liver and circulation and plays a protective role in hepatic lipid accumulation. To investigate the mechanism by which FGF21 protects against liver lipid accumulation under protein malnutrition, we determined whether upregulated FGF21 promotes the thermogenesis or secretion of very-low-density lipoprotein (VLDL)-triacylglycerol (TAG). The results showed that protein malnutrition decreased VLDL-TAG secretion, but the upregulation of FGF21 did not oppose this effect. In addition, protein malnutrition increased expression of the thermogenic gene uncoupling protein 1 in inguinal white adipose and brown adipose tissue in an FGF21-dependent manner. However, surgically removing inguinal white adipose tissue did not affect liver triglyceride levels in protein-malnourished mice. These data suggest that FGF21 stimulates thermogenesis under protein malnutrition, but this is not the causative factor underlying the protective role of FGF21 against liver lipid accumulation.

Amino acid losses are lower during pre-dilution on-line HDF than HD of the same Kt/V for urea.

In this study, we investigated differences in amino acid losses between HD and pre-dilution on-line HDF with equal Kt/V for urea to determine which modality removes less amino acids from extravascular pools and ensures better nutrition. The subjects were patients receiving pre-dilution on-line HDF (n = 10) or HD (n = 10) at this hospital. Dialysis time was 4 h for all patients. In patients on HD, the blood flow rate was 200 mL/min and the dialysate flow rate was 463 ± 29.3 mL/min. In patients on pre-dilution on-line HDF, the blood flow rate was 240 ± 20 mL/min, the dialysate flow rate was 565.0 ± 42.5 mL/min, and the substitution flow rate (substitution volume) was 252.8 ± 26.4 mL/min (57.0 ± 6.0 L). Kt/V for urea was comparable between patients on HD and patients on pre-dilution on-line HDF (1.46 ± 0.25 vs. 1.46 ± 0.31). Amino acid loss and clear space were evaluated. Patients on pre-dilution on-line HDF lost significantly less glutamine and arginine (p < 0.01 and p = 0.032) and significantly less nonessential amino acids (NEAAs) than patients on HD (p = 0.013). They also had significantly lower clear space of total amino acids (TAAs), NEAAs, essential amino acids (EAAs), and branched-chain amino acids (BCAAs) than patients on HD (Total AA p = 0.019, NEAA p = 0.018, EAA p = 0.024, BCAA p = 0.042). When Kt/V for urea is equal, pre-dilution on-line HDF ensures better nutrition than does HD.

Restless legs syndrome effectively treated with constant-pressure predilution online hemodiafiltration.

BACKGROUND: We encountered a case of unstable predilution online HDF due to elevated transmembrane pressure (TMP) when performing constant-speed predilution online hemodiafiltration (HDF) as treatment for restless legs syndrome (RLS) in a dialysis patient. We report the effectiveness of incorporating a newly developed constant-pressure predilution online HDF system as a preventive measure against unstable online HDF and frequent adjustment of settings when treating dialysis patients with RLS. CASE PRESENTATION: A 55-year-old man had suffered from RLS and been undergoing constant-speed online HDF with 45 L target predilution and an ABH-21P hemodiafilter. The symptoms of RLS rated 10 on the International Restless Legs Syndrome Rating Scale (IRLS). The α1-microglobulin (α1-MG) removal rate was only 27.8%, so the hemodiafilter was subsequently replaced with a PEPA hemodiafilter. However, episodes of elevated TMP exceeding 250 mmHg occurred frequently after the replacement and were managed by reducing dialysate flow rate. Therefore, we incorporated a constant-pressure predilution online HDF that maintains TMP below 200 mmHg. The amount of replacement was maintained at approximately 43.5 ± 6.98 L and the α1-MG removal rate was 39.5%, with no need to manually reduce the flow rate. The Alb leakage in dialysate waste was 7.9 g. The patient has maintained an IRLS rating of 0 with no RLS symptoms for the past 4 years. CONCLUSIONS: Using the constant-pressure mode enabled achieved the clinical endpoint, namely, resolution of RLS with no need to manually reduce the flow rate.

Effects of pharmacological inhibition of plasminogen binding on liver regeneration in rats.

The fibrinolysis system is thought to play an important role in liver regeneration. We previously found that plasminogen (Plg) is localized to the cell surface of regenerating liver tissue as well as proliferating hepatocytes in vitro. Here, we investigated the significance of Plg binding to the cell surface during liver regeneration. Pre-administration of tranexamic acid (TXA), which is a competitive inhibitor of Plg binding, to hepatectomized rats mildly delayed restoration of liver weight in vivo. Although binding of Plg to the cell membrane decreased following TXA administration, TXA showed little effect on hepatocyte proliferation in rats. We also discovered that Plg treatment did not stimulate proliferation of primary rat hepatocytes in vitro. These results suggest that Plg/plasmin potentiates liver regeneration via a pathway distinct from those through which hepatocyte proliferation is stimulated.

New diagnostic criteria and operative strategy for cesarean scar syndrome: Endoscopic repair for secondary infertility caused by cesarean scar defect.

AIM: The aim of the present study was to assess the efficacy of endoscopic repair for secondary infertility caused by post-cesarean scar defect (PCSD). Our investigation focused on the validity of new diagnostic criteria and selection methods. MATERIAL AND METHODS: The subjects were 22 women with secondary infertility due to PCSD with retention of bloody fluid in the uterine cavity. Women with a residual myometrial thickness of ≥ 2.5 mm and an anteflexed or straight uterus underwent hysteroscopic surgery, while all others underwent laparoscopic repair. Hysteroscopic surgery involved resection and coagulation of scarred areas, whereas laparoscopic surgery involved removal of scarred areas combined with hysteroscopy, followed by resuturing. RESULTS: Fourteen of the 22 women (63.6%) who were followed up for ≥ 1 year after surgery achieved pregnancy. Pregnancies occurred in all four women (100%) who underwent hysteroscopic surgery and in 10 of the 18 women (55.6%) who underwent laparoscopic surgery. Three out of four women who underwent hysteroscopic surgery had term deliveries. Among the women who underwent laparoscopic surgery, five had term deliveries. No cases of uterine rupture were experienced, and the delivery method was cesarean section in all cases. CONCLUSION: We propose that infertility associated with PCSD, cesarean scar syndrome, is caused by the retention of bloody fluid in the uterine cavity and scarring. Endoscopic treatment, such as hysteroscopy or laparoscopy, was effective for cesarean scar syndrome.

Diallyl trisulfide induces apoptosis in Jurkat cells by the modification of cysteine residues in thioredoxin.

We reported the regulation of protein function by oxidative modification of the specific cysteine residue(s) by diallyl trisulfide (DATS). In this study, we examined if DATS modifies the cysteine residue of thioredoxin (Trx) by urea-polyacryl amide gel electrophoresis. DATS modified two specific cysteine residues in Trx and this oxidative modification of cysteine residues would be sole causative of the apoptosis induced by DATS in leukemic cells.

Identification of molecular target of diallyl trisulfide in leukemic cells.

To identify the molecular target of diallyl trisulfide (DATS) in human leukemic cell line U937, we examined modification of thiol group(s) of cellular proteins by the redox 2D PAGE. A unique protein spot appeared by DATS treatment was identified to be heat shock protein 27 (HSP27). Hsp27 is suggested to be one of the molecular target of DATS in U937.

Development of marine biodiversity database (BISMaL) to enable estimations past habitat conditions for marine life in the northwestern Pacific.

Global activities involving the collection of marine biodiversity information have provided a large amount of biological observation records that cover various spatiotemporal areas. To predict biological responses or distribution changes in response to environmental changes by using these observation records, it is essential to analyze not only the current marine physicochemical environmental conditions but also the past conditions when the organisms were observed. We developed a new function to estimate the past marine environmental conditions for the observation records in our marine biodiversity database (Biological Information System for Marine Life: BISMaL) and examine whether the database can reliably estimate thermal habitats for both benthic and planktonic marine organisms. For the benthic squat lobster Shinkaia crosnieri, the estimated and observed in situ temperatures were similar to each other. For the planktonic chaetognaths Krohnitta pacifica and K. subtilis, the estimated temperatures showed clear seasonal changes specific to their distribution areas. These results indicated that BISMaL can reliably provide past habitat conditions regardless of planktonic or benthic lifestyles. BISMaL, which provides both biological observations and estimated past environmental conditions through web services, could lower the barrier to data access and use and make data-driven science available not only for data scientists but also for various marine scientists, such as taxonomists, ecologists and field scientists. Database URL:  https://www.godac.jamstec.go.jp/bismal/e/.

Two oxytocin analogs, N-(p-fluorobenzyl) glycine and N-(3-hydroxypropyl) glycine, induce uterine contractions ex vivo in ways that differ from that of oxytocin.

Contraction of the uterus is critical for parturient processes. Insufficient uterine tone, resulting in atony, can potentiate postpartum hemorrhage; thus, it is a major risk factor and is the main cause of maternity-related deaths worldwide. Oxytocin (OT) is recommended for use in combination with other uterotonics for cases of refractory uterine atony. However, as the effect of OT dose on uterine contraction and control of blood loss during cesarean delivery for labor arrest are highly associated with side effects, small amounts of uterotonics may be used to elicit rapid and superior uterine contraction. We have previously synthesized OT analogs 2 and 5, prolines at the 7th positions of which were replaced with N-(p-fluorobenzyl) glycine [thus, compound 2 is now called fluorobenzyl (FBOT)] or N-(3-hydroxypropyl) glycine [compound 5 is now called hydroxypropyl (HPOT)], which exhibited highly potent binding affinities for human OT receptors in vitro. In this study, we measured the ex vivo effects of FBOT and HPOT on contractions of uteri isolated from human cesarean delivery samples and virgin female mice. We evaluated the potency and efficacy of the analogs on uterine contraction, additivity with OT, and the ability to overcome the effects of atosiban, an OT antagonist. In human samples, the potency rank judged by the calculated EC50 (pM) was as follows: HPOT (189) > FBOT (556) > OT (5,340) > carbetocin (12,090). The calculated Emax was 86% for FBOT and 75% for HPOT (100%). Recovery from atosiban inhibition after HPOT treatment was as potent as that after OT treatment. HPOT showed additivity with OT. FBOT (56 pM) was found to be the strongest agonist in virgin mouse uterus. HPOT and FBOT demonstrated high potency and partial agonist efficacy in the human uterus. These results suggested that HPOT and FBOT are highly uterotonic for the human uterus and performed better than OT, indicating that they may prevent postpartum hemorrhage.

Lipoprotein lipase is a novel amyloid beta (Abeta)-binding protein that promotes glycosaminoglycan-dependent cellular uptake of Abeta in astrocytes.

Lipoprotein lipase (LPL) is a member of a lipase family known to hydrolyze triglyceride molecules in plasma lipoprotein particles. LPL also plays a role in the binding of lipoprotein particles to cell-surface molecules, including sulfated glycosaminoglycans (GAGs). LPL is predominantly expressed in adipose and muscle but is also highly expressed in the brain where its specific roles are unknown. It has been shown that LPL is colocalized with senile plaques in Alzheimer disease (AD) brains, and its mutations are associated with the severity of AD pathophysiological features. In this study, we identified a novel function of LPL; that is, LPL binds to amyloid ß protein (Aß) and promotes cell-surface association and uptake of Aß in mouse primary astrocytes. The internalized Aß was degraded within 12 h, mainly in a lysosomal pathway. We also found that sulfated GAGs were involved in the LPL-mediated cellular uptake of Aß. Apolipoprotein E was dispensable in the LPL-mediated uptake of Aß. Our findings indicate that LPL is a novel Aß-binding protein promoting cellular uptake and subsequent degradation of Aß.

Apolipoprotein E regulates the integrity of tight junctions in an isoform-dependent manner in an in vitro blood-brain barrier model.

Apolipoprotein E (apoE) is a major apolipoprotein in the brain. The ε4 allele of apoE is a major risk factor for Alzheimer disease, and apoE deficiency in mice leads to blood-brain barrier (BBB) leakage. However, the effect of apoE isoforms on BBB properties are as yet unknown. Here, using an in vitro BBB model consisting of brain endothelial cells and pericytes prepared from wild-type (WT) mice, and primary astrocytes prepared from human apoE3- and apoE4-knock-in mice, we show that the barrier function of tight junctions (TJs) was impaired when the BBB was reconstituted with primary astrocytes from apoE4-knock-in mice (apoE4-BBB model). The phosphorylation of occludin at Thr residues and the activation of protein kinase C (PKC)η in mBECs were attenuated in the apoE4-BBB model compared with those in the apoE3-BBB model. The differential effects of apoE isoforms on the activation of PKCη, the phosphorylation of occludin at Thr residues, and TJ integrity were abolished following the treatment with an anti-low density lipoprotein receptor-related protein 1 (LRP1) antibody or a LRP1 antagonist receptor-associated protein. Consistent with the results of in vitro studies, BBB permeability was higher in apoE4-knock-in mice than in apoE3-knock-in mice. Our studies provide evidence that TJ integrity in BBB is regulated by apoE in an isoform-dependent manner.

Advanced glycation endproducts stimulate renal epithelial cells to release chemokines that recruit macrophages, leading to renal fibrosis.

Diabetic nephropathy is a major complication of diabetes and tubulointerstitial fibrosis is one of its manifestations. This study aimed to clarify the pathogenicity of advanced glycation endproducts (AGEs) toward NRK-52E, a tubular epithelial cell line. The AGE-exposed cells significantly increased gene expression of transforming growth factor beta, plasminogen activator inhibitor-1, and tissue transglutaminase, and a medium conditioned by them showed strong potential to recruit macrophages, partly through a chemokine, monocyte chemoattractant protein-1. Albumin denatured by maintenance at 37 °C for 120 d exhibited similar activities, but they were lower than those of the AGEs. Thus, AGEs generated in diabetic patients might exacerbate fibrosis in the kidneys directly through renal epithelial cell stimulation, and indirectly by recruitment of macrophages.

Cinnamon extract improves abnormalities in glucose tolerance by decreasing Acyl-CoA synthetase long-chain family 1 expression in adipocytes.

We previously demonstrated that cinnamon extract (CE) alleviates streptozotocin-induced type 1 diabetes in rats. The present study aimed to elucidate the detailed molecular target of cinnamon in cultured adipocytes and epididymal adipose tissue of type 2 diabetes model mice. Two-dimensional gel electrophoresis was employed to determine the molecular target of cinnamon in adipocytes. The function of Acyl-CoA synthetase long-chain family-1 (ACSL1), a molecular target of cinnamon that was identified in this study, was further investigated in 3T3-L1 adipocytes using specific inhibitors. Type 2 diabetes model mice (KK-Ay/TaJcl) were used to investigate the effect of CE on glucose tolerance, ACSL1 expression, and related signal molecules in vivo. CE decreased ACSL1 mRNA and protein expression in 3T3-L1 adipocytes but increased glucose uptake and AMPK signaling activation; moreover, a similar effect was observed with an ACSL1 inhibitor. CE improved glucose tolerance and downregulated ACSL1 in mice adipose tissue in vivo. ACSL1 was demonstrated as a molecular target of CE in type 2 diabetes both in a cell culture system and diabetic mouse model.