Female ICR Mice Research Articles

Climbing behavior by mice as an endpoint for preclinical assessment of drug effects in the absence and presence of pain.

This study evaluated climbing in mice as a tool to assess the expression and treatment of pain-related behavioral depression in male and female ICR mice. Mice were videotaped during 10-min sessions in a vertical plexiglass cylinder with wire mesh walls, and "Time Climbing" was scored by observers blind to treatments. Initial validation studies demonstrated that baseline climbing was stable across repeated days of testing and depressed by intraperitoneal injection of dilute lactic acid (IP acid) as an acute pain stimulus. Additionally, IP acid-induced depression of climbing was blocked by the positive-control non-steroidal anti-inflammatory drug (NSAID) ketoprofen but not by the negative control kappa opioid receptor agonist U69593. Subsequent studies examined effects of single-molecule opioids (fentanyl, buprenorphine, naltrexone) and of fixed-proportion fentanyl/naltrexone mixtures (10:1, 3.2:1, and 1:1) that vary in their efficacy at the mu opioid receptor (MOR). Opioids administered alone produced a dose- and efficacy-dependent decrease in climbing, and fentanyl/naltrexone-mixture data indicated that climbing in mice is especially sensitive to disruption by even low-efficacy MOR activation. Opioids administered as a pretreatment to IP acid failed to block IP acid-induced depression of climbing. Taken together, these findings support the utility of climbing in mice as an endpoint to evaluate candidate-analgesic effectiveness both to (a) produce undesirable behavioral disruption when the test drug is administered alone, and (b) produce a therapeutic blockade of pain-related behavioral depression. The failure of MOR agonists to block IP acid-induced depression of climbing likely reflects the high sensitivity of climbing to disruption by MOR agonists.

Exposure to PCB126 during the nursing period reversibly impacts early-life glucose tolerance.

Polychlorinated biphenyls (PCBs) are persistent environmental organic pollutants known to have detrimental health effects. Using a mouse model, we previously demonstrated that PCB126 exposure before and during pregnancy and throughout the perinatal period adversely affected offspring glucose tolerance and/or body composition profiles. The purpose of this study was to investigate the glucose tolerance and body composition of offspring born to dams exposed to PCB126 during the nursing period only. Female ICR mice were bred, and half of the dams were exposed to either vehicle (safflower oil) or 1 µmole PCB126 per kg of body weight via oral gavage on postnatal days (PND) 3, 10, and 17 (n = 9 per group). Offspring body weight, lean and fat mass, and glucose tolerance were recorded every three weeks. PCB126 treatment did not alter dam nor offspring body weight (p > 0.05). PCB126-exposed male and female offspring displayed normal body composition (p > 0.05) relative to vehicle-exposed offspring. However, both male and female offspring that were exposed to PCB126 during the nursing period had significantly impaired glucose tolerance at 3 and 9 weeks of age (p < 0.05). At 6 and 12 weeks of age, no impairments in glucose tolerance existed in offspring (p > 0.05). Our current study demonstrates that exposure to PCB126 through the mother's milk does not affect short- or long-term body composition but impairs glucose tolerance in the short-term.

Establishment of an overactive bladder model in mice

BackgroundOveractive bladder (OAB) is a syndrome characterized by symptoms of urinary urgency, often accompanied by frequent urination and nocturia or urge incontinence.MethodsTwenty female ICR mice were randomly divided into pBOO (partial bladder outlet obstruction) and control groups. The mouse OAB model was constructed by ligating the bladder outlet. Eight weeks after the operation, the methods of voiding spot on paper (VSOP), isolated detrusor muscle, and HE staining were used for analysis and research.ResultsAfter the operation, two mice in the experimental and one in control died, and one in the control groups had an abnormal bladder size, so it was excluded from the statistical analysis. Eight weeks after the operation, there was an insignificant difference (P = 0.15) in the body weight of mice in the pBOO (26.54 ± 2.62 g) and the control group (24.84 ± 1.76 g). The number of urinations in 12 h was significantly higher (P < 0.001) in the pBOO (7.63 ± 1.19) than in the control group (4.13 ± 0.99). Also, the 12-h urine volume of pBOO (1491.23 ± 94.72 μL) was significantly greater (P = 0.006) than that of the control group (1344.86 ± 88.17 μL). The isolated bladder of the pBOO mice was significantly heavier than that in the control group (53.16 ± 1.79 mg vs. 24.54 ± 1.80 mg, P < 0.001), the horizontal and vertical length of the bladder in pBOO group were larger than those in the control group (P < 0.001). The detrusor thickness of pBOO group (357.50 ± 11.88 µm) was significantly thicker than that of control group (258.52 ± 17.22 µm, P < 0.001), and the isolated muscle strip was more sensitive to carbachol stimulation. According to HE staining, the bladder wall of the pBOO mice was significantly thickened.ConclusionsA pBOO-mediated mouse OAB model was successfully established by ligating the bladder outlet.

Determination of the synergistic anti-influenza effect of Huangqin Su tablet and Oseltamivir and investigation of mechanism of the tablet based on gut microbiota and network pharmacology

Huangqin Su (HQS) tablet is mainly composed of baicalein which has been evaluated for its ability to inhibit influenza. The present study aimed to investigate the effect of HQS and oseltamivir phosphate (OS) (single or combination therapy) on influenza-induced acute pneumonia in male and female ICR mice. The regulatory effect of HQS on gut microbiota was also studied by using 16 s rDNA sequencing, and the targets and mechanisms of HQS against influenza were comprehensively analyzed by network pharmacology. Pharmacodynamic results, including lung index and pathological changes, showed that HQS exhibited significant anti-influenza efficacy and could improve the efficacy of low-dose OS (P < 0.05 and P < 0.01, respectively). The results of 16 s rDNA sequencing revealed that HQS modulated the gut microbiota and remarkably enriched the abundance of Lactobacillus. The findings of network pharmacology research suggested that the anti-influenza mechanism of HQS was related to TLRs, MAPK, and other signal transduction pathways. Taken together, this study identified the possibility of the combined use of HQS and OS and demonstrated the role of HQS in modulating the gut microbiota of mice against influenza. Network pharmacology studies also suggested that the anti-influenza effect of HQS was related to TLRs, MAPK, TNF, and other signaling pathways.

High fat diet induces obesity, alters eating pattern and disrupts corticosterone circadian rhythms in female ICR mice.

Circadian, metabolic, and reproductive systems are inter-regulated. Excessive fatness and circadian disruption alter normal physiology and the endocrine milieu, including cortisol, the primary stress hormone. Our aim was to determine the effect feeding a high fat diet to female ICR mice had on diurnal feeding pattern, weight gain, body composition, hair corticosterone levels and circadian patterns of fecal corticosterone. Prepubertal (~35d of age) ICR mice were assigned to control (CON; 10% fat) or high fat (HF; 60% fat) diet and fed for 4 wk to achieve obesity under 12h light and 12h of dark. Feed intake was measured twice daily to determine diurnal intake. Mice were weighed weekly. After 4 wk on diets hair was collected to measure corticosterone, crown-rump length was measured to calculate body mass index (BMI), and body composition was measured with EchoMRI to determine percent fat. HF mice weighed more (P<0.05) after week two, BMI and percent body fat was greater (P<0.05) in HF than CON at the end of wk 4. HF mice consumed more during the day (P<0.05) than CON mice after 1 week on diets. Hair corticosterone was higher in HF mice than in CON (P<0.05). Fecal circadian sampling over 48hr demonstrated that HF mice had elevated basal corticosterone, attenuated circadian rhythms, and a shift in amplitude. High fat feeding for diet induced obesity alters circadian eating pattern and corticosterone rhythms, indicating a need to consider the impact of circadian system disruption on reproductive competence.

Benincasa hispida Extract Prevents Ovariectomy-Induced Osteoporosis in Female ICR Mice

With the increase in bone metabolic diseases owing to the aging of the global population, interest in functional food ingredients for improving bone health is increasing. This study aimed to determine the anti-osteoporosis effect of Benincasa hispida extract (BHE, HR1901-W) and 2-furoic acid in ovariectomy (OVX)-induced osteoporosis in female ICR mice. Thirty-five female ICR mice underwent OVX or sham operation and were randomized into seven groups of five animals as follows: normal, sham, OVX, OVX with genistein (10 mg/kg), 2-furoic acid (20 mg/kg), LBHE (100 mg/kg), and HBHE (200 mg/kg). After an 8-week treatment period, femur and blood samples were collected from mice. Bone density and bone formation markers were significantly recovered in the 2-furoic acid and HBHE supplementation groups compared with those in the OVX group. In addition, bone resorption markers were increased in OVX mice, whereas they were significantly decreased in the OVX + 2-furoic acid and HBHE supplementation groups. This study suggests that BHE supplementation prevents bone resorption and promotes bone formation in OVX mice. These findings indicate that BHE could be used as a promising natural means to prevent OVX-induced osteoporosis and bone metabolic diseases.

Evaluation of Clastogenic/Aneugenic Damage Using the FISH Micronucleus Assay in Mice Exposed to Chromium (VI).

Exposure to chromium (VI) [Cr(VI)] has been postulated to be associated with the induction of cancer. In vivo studies utilizing biomarkers of genotoxic damage could aid in elucidating the mechanisms underlying the genotoxic effects of Cr(VI) and their relationship with carcinogenesis. In this study, the origin (clastogenic and/or aneugenic damage) and kinetics of micronuclei (MN) induced by Cr(VI) were investigated. Hsd:ICR female mice were divided into groups of five individuals each. MN kinetics were measured in groups treated with 20 or 25 mg/kg CrO3 intraperitoneally using acridine orange-coated slides in peripheral blood obtained from the caudal vein 0, 12, 24, 36, 48, 60, and 72 h after treatment. Whereas identification of MN with centromeric DNA (MNK+) was measured at the dose of 20 mg/kg of CrO3, using fluorescence in situ hybridization (FISH) with a centromere-specific probe in peripheral blood obtained at 0, 12, and 48 h after treatment. Control groups were administered vehicle only. Total MN were quantified and the clastogenic/aneugenic effects of Cr(VI) were evaluated based on the proportion of MNK+ versus micronuclei without centromeric DNA (MNK-). There was a significant increase in MN frequencies beginning at 12 h in the Cr(VI)-treated groups demonstrating its genotoxicity. When calculating the MNK+ as a percentage of the total MN, the increase was significant beginning 12 h after treatment. The fact that the MNK+ and MNK- were observed at both evaluation times corroborates Cr(VI) as a genotoxic agent and demonstrates that both clastogenic and aneugenic damages are involved in the formation of MN.

PS-B05-9: ADVANCED MATERNAL AGED MOUSE MODEL SHOWED FETAL GROWTH RESTRICTION AND WOULD BE A MODEL FOR DOHAD

Backgrounds: The number of advance-aged pregnancies, defined as those over 35 years of age, has been increasing in developed countries due to tendency to marry later with women's social advancement and assisted reproductive technology. Furthermore, in Japan, the incidence of hypertensive disorders of pregnancy over 40 years of age is as high as approximately 8%, which have a high rate of premature births and low birth weight, and fewer nephrons. Recently, based on the developmental origins of health and disease (DOHaD) hypothesis, it has been recognized that the risk of developing hypertension, proteinuria and renal failure in the future is increased in aged pregnancies. However, the detailed mechanism is uncertain. Methods: Female ICR mice of 6 to 10 months (Advanced maternal age, AMA group) and of 10 to 12 weeks (Control; CON group) were mated with males of 10 to 16 weeks. Blood pressure during pregnancy on day 7 and 14 was measured in both groups (n = 4, n = 4). The pups born in both groups (AMA, n = 4–13; CON, n = 10–16) were weighed at postnatal day 0, a week, 4 weeks, and 8 weeks of age. Blood pressure (n = 7, n = 3) at 8 weeks of age was measured using the tail-cuff technique. The results were compared using the Mann-Whitney U test. Results: There was no difference in blood pressure during pregnancy on day 7 and day 14 in both groups. In the AMA mouse model, the weights of postnatal day 0 pups were significantly lower (AMA vs. CON, 1.66 ± 0.22 g vs. 1.9 ± 0.22 g, P = 0.006). However, no difference was found between the two groups at a week and 4 weeks. Conversely, the body weights of the AMA model were significantly higher at 8 weeks of age (AMA vs. CON, 47.7 ± 3.43 g vs. 45.4 ± 2.8 g, P = 0.041) and the systolic blood pressure in the AMA group tended to be higher. Discussion/Conclusion: In the present study, the AMA mouse model induced fetal growth restriction and subsequently showed catch-up growth. Furthermore, their pups’ weights were significantly higher and elevated blood pressure was observed at adult age. While previous animal models of DOHaD have been mainly based on studies of dietary restriction, chemical exposure, and preeclampsia, this model showed similar symptoms to DOHaD models.

Maternal circadian disruption before pregnancy impairs the ovarian function of female offspring in mice

Circadian disturbance brought on by shift employment, nighttime light pollution, and other factors is quite prevalent in contemporary culture. However, the effect of maternal circadian disruption before pregnancy on the reproduction of offspring in mice requires further research. Herein, we exposed female ICR mice to constant light to establish a model of preconceptional circadian disruption and then checked the ovarian function of female offspring (named the CLE group below). Our results revealed obesity, abnormal lipid metabolism and earlier puberty onset in the CLE group. Additionally, impaired ovarian follicle development, oocyte quality and preimplantation embryo development were shown in the CLE group. Moreover, the expression levels of Gnrh1 in the hypothalamus and Cyp17a1, Bmper, Bdnf and Lyve1 in ovaries, as well as circadian clock genes, including Clock, Cry1, Nr1d2 and Per2, were significantly downregulated in the CLE group. Mechanistically, immune responses, including the interleukin-17 (IL-17) signalling pathway, cytokine–cytokine receptor interaction and the chemokine signalling pathway, were altered in the CLE group, which may be responsible for the damaged ovarian function.

Sub-acute Oral Toxicity Evaluation of Philippine Bignay (Antidesma bunius (L.) Spreng cv. ‘Kalabaw’) Fruit in ICR Mice

This study evaluated the safety of Bignay (Antidesma bunius (L.) Spreng cv. ‘Kalabaw’) fruits through a sub-acute oral toxicity test in male and female ICR mice. The evaluation was done through oral gavage of the aqueous fruit sample at 2,000 mg kg-1 body weight dose for 28 d. Results showed that there were no significant changes in feed and water intake of male and female ICR mice as compared to the control groups that received distilled water (p &lt; 0.05). Blood chemistry and hematology values for all mice groups were comparable to the controls and published values. All mice groups also had appreciable body weight gain (10.89% – 21.69%) with zero morbidity and mortality. Lastly, the gross and microscopic examination of the brain, heart, lungs, GI tract, liver, spleen, and kidneys showed normal architecture and histology, suggesting that the fruits did not induce any morphological abnormalities. Results showed that Bignay (cv. ‘Kalabaw’) fruits are safe for consumption with an LD50 of greater than 2,000 mg kg-1 body weight.

Acute and Subacute Toxicity of Fluorescent Gold Nanoclusters Conjugated with α-Lipoic Acid.

Fluorescent gold nanoclusters conjugated with α-lipoic acid (FANC) is a promising biocompatible fluorescent nanomaterial with a high potential for drug development. However, there is still no FANC-related research on toxicology, which is very important for future research and the development of healthy food supplements or drugs. This study uses oral administration of FANC to determine the most appropriate dose range in ICR mice for further evaluation. The in vivo acute and subacute toxicity study was conducted by oral administration of FANC to male and female ICR mice. Animal survival, body weight, daily food consumption, hematological profile, organ coefficient, serum biochemistry profile, and histopathological changes were analyzed. FANC did not show any form of morbidity or mortality at acute and subacute toxicity in both male and female ICR mice. Animal behavior, daily food consumption, hematological profile, organ coefficient, and histopathology showed no treatment-related malignant changes at single and repeated doses. Furthermore, serum glutamic-oxaloacetic transaminase (GOT), glutamic-pyruvic transaminase (GPT), lactate dehydrogenase (LDH), blood urea nitrogen (BUN), and creatinine (CRE) levels showed no significant malignant changes, which indicated that FANC does not cause liver and renal damage. The only change observed in this study was the change in body weight. The body weight of the FANC-treated group was slightly decreased in female mice but increased in male mice; however, the body weight decreases were below the threshold of concern, and there was no dose-response effect. In conclusion, no observed adverse effect level (NOAEL) in repeated doses was considered in 20 μM/100 μL/25 g male and female ICR mice.

The Corticosterone-Glucocorticoid Receptor-AP1/CREB Axis Inhibits the Luteinizing Hormone Receptor Expression in Mouse Granulosa Cells.

Under stress conditions, luteinizing hormone (LH)-mediated ovulation is inhibited, resulting in insufficient oocyte production and excretion during follicular development. When the body is stressed, a large amount of corticosterone (CORT) is generated, which will lead to a disorder of the body’s endocrine system and damage to the body. Our previous work showed that CORT can block follicular development in mice. Since LH acts through binding with the luteinizing hormone receptor (Lhcgr), the present study aimed to investigate whether and how corticosterone (CORT) influences Lhcgr expression in mouse ovarian granulosa cells (GCs). For this purpose, three-week-old ICR female mice were injected intraperitoneally with pregnant mare serum gonadotropin (PMSG). In addition, the treatment group was injected with CORT (1 mg/mouse) at intervals of 8 h and the control group was injected with the same volume of methyl sulfoxide (DMSO). GCs were collected at 24 h, 48 h, and 55 h after PMSG injection. For in vitro experiments, the mouse GCs obtained from healthy follicles were treated with CORT alone, or together with inhibitors against the glucocorticoid receptor (Nr3c1). The results showed that the CORT caused a downregulation of Lhcgr expression in GCs, which was accompanied by impaired cell viability. Moreover, the effect of the CORT was mediated by binding to its receptor (Nr3c1) in GCs. Further investigation revealed that Nr3c1 might regulate the transcription of Lhcgr through inhibiting the expression of Lhcgr transcription factors, including AP1 and Creb. Taken together, our findings suggested a possible mechanism of CORT-induced anovulation involving the inhibition of Lhcgr expression in GCs by the CORT–Nr3c1–AP1/Creb axis.

Co-Expression of Pig IL-2 and Fusion Bovine Cathelicidin Gene by Recombinant Plasmids in Yeast and Their Promotion of Mouse Antibacterial Defense.

Simple Summary1. Oral inoculation with recombinant yeast co-expressing pig IL-2 and fused bovine cathelicidin (FBC) significantly enhanced systemic immunity in mice. 2. The inoculated mice displayed stronger resistance against E. coli or S. aureus infection. 3. Recombinant yeast co-expressing PIL-2 and FBC could facilitate the development of new immunopotentiator to prevent the infection of antibiotic-resistant bacteria.In order to develop an effective and safe immunomodulator to enhance the antimicrobial bioactivity and immunity of animals against infectious bacterial diseases, a recombinant plasmid pGAPZαA-IL2-B co-expressing pig interleukin-2 (PIL-2) and fused bovine cathelicidin (FBC) genes were constructed using the 2A self-cleavage technique. After being expressed in Pichia pastoris strain SMD1168, the recombinant yeast was administered orally to 5-week-old female ICR mice. The control mice were similarly dosed with P. pastoris with a blank plasmid or FBC recombinant plasmid alone. At 28 days post-treatment, the mice were challenged intraperitoneally with virulent strains of either E. coli or S. aureus. Compared with the control groups, the mice that received recombinant yeast co-expressing PIL-2/FBC manifested significant increases in the number of leukocytes, CD4+ and CD8+ T cells, IgG, and the gene expressions of TLRs(TLR1,4,6,9), antimicrobial peptides(CRP4 and CRAMP) and cytokines (IL-2, 4, 6, 7, 12, 15, 23, IFN-γ, and TNF-α) in the blood. Furthermore, the treated mice displayed significantly higher survival than the other two control groups after the challenge. These results suggest that the antimicrobial activity and immunity of animals can be effectively enhanced by the in vivo co-expression of IL-2 and the FBS gene, which can facilitate the development of new immunopotentiation molecules to overcome the infection of antibiotic-resistant bacteria.

Anti-Osteoporosis Effects of the Fruit of Sea Buckthorn (Hippophae rhamnoides) through Promotion of Osteogenic Differentiation in Ovariectomized Mice.

The fruit of Hippophae rhamnoides has been widely used for medicinal purposes because of its anti-inflammatory, antioxidant, antiplatelet, and antimicrobial effects. Since there are no clear reports on the therapeutic efficacy of H. rhamnoides in osteoporosis, this study aimed to confirm the potential use of H. rhamnoides for the treatment of osteoporosis through its osteogenic differentiation-promoting effect in ovariectomized mice. Through an in vitro study, we compared the effects of the EtOH extract of H. rhamnoides fruits (EHRF) on the differentiation of C3H10T1/2, a mouse mesenchymal stem cell line, into osteoblasts based on alkaline phosphatase (ALP) staining and the relative expression of osteogenesis-related mRNAs. The EHRF significantly stimulated the differentiation of mesenchymal stem cells into osteoblasts and showed 7.5 times (* p < 0.05) higher osteogenesis than in the untreated control. A solvent fractionation process of EHRF showed that the hexane-soluble fraction (HRH) showed 10.4 times (** p < 0.01) higher osteogenesis than in the untreated control. Among the subfractions derived from the active HRH by preparative HPLC fractionation, HRHF4 showed 7.5 times (* p < 0.05) higher osteogenesis than in the untreated naïve cells, and HRH and HRHF4 fractions showed 22.6 times (*** p < 0.001) stronger osteogenesis activity than in the negative control. Osteoporosis was induced by excision of both ovaries in 9-week-old female ICR mice for in vivo analysis, and two active fractions, HRH and HRHF4, were administered orally for three months. During the oral administration period, body weight was measured weekly, and bone mineral density (BMD) and body fat density were measured simultaneously using a DEXA machine once a month. In particular, during the in vivo study, the average BMD of the ovariectomized group decreased by 0.0009 g/cm2, whereas the average BMD of the HRH intake group increased by 0.0033 g/cm2 (* p < 0.05) and that of the HRHF4 intake group increased by 0.0059 g/cm2 (** p < 0.01). The HRH and HRHF4 intake groups significantly recovered the mRNA and protein expression of osteogenic genes, including ALP, Osteopontin, Runx2, and Osterix, in the osteoporosis mouse tibia. These findings suggest that the active fractions of H. rhamnoides fruit significantly promoted osteoblast differentiation in mesenchymal stem cells and increased osteogenic gene expression, resulting in an improvement in bone mineral density in the osteoporosis mouse model. Taken together, H. rhamnoides fruits are promising candidates for the prevention and treatment of osteoporosis.

Integration of pharmacodynamics and metabolomics reveals the therapeutic effects of 6-acetylacteoside on ovariectomy-induced osteoporosis mice

Background6-acetylacteoside (6-AA) is a phenylethanoid glycoside isolated from Cistanche deserticola which had been previously proven to possess anti-osteoporotic activity previously. Currently, it is still unknown whether 6-AA plays a crucial role on the anti-osteoporotic effects of C. deserticola. PurposeTo elucidate the therapeutic effect and mechanism of 6-AA on osteoporosis by employing an ovariectomized mouse model in vivo and RAW264.7 cells in vitro. MethodsSixty female ICR mice were randomly assigned into six groups: sham-operated control group (SHAM, vehicle), ovariectomized model group (OVX, vehicle), positive group (EV, 1 mg/kg/day of estradiol valerate), low dosage (10 mg/kg/day of 6-AA), medium dosage (20 mg/kg/day of 6-AA) and high dosage (40 mg/kg/day of 6-AA) treatment groups. All substances were administered daily by intragastric gavage. After 12 weeks of intervention, trabecular bone microarchitecture was estimated and bone biomechanics were determined. Bone formation and resorption factors were determined by using the corresponding Elisa kits. The related proteins and metabolites were estimated by using western-blot and metabolomics techniques. ResultsOVX mice demonstrated significant atrophy of the uterine and vagina, declined biomechanical parameters such as flexural strength and maximum load, deteriorated trabecular bone microarchitecture such as decreased BMD, BMC, TMC, TMD, BVF, Tb. N, and Tb. Th and increased Tb. Sp, as well as increased bone resorption factors such as TRAP, cathepsin K, and DPD, all after 12 weeks of ovariectomy operation. Following administration of 6-AA to OVX mice, parameters related to the bone microarchitecture, bone resorption activities as well as biomechanical properties were all significantly improved. Meanwhile, the levels of NF-κB, NFATc1, RANK, RANKL and TRAF6 were significantly downregulated, while OPG, PI3K and AKT were upregulated after 6-AA intervention. This indicates that, 6-AA could prevent bone resorption by regulating the RANKL/RANK/OPG mediated NF-κB and PI3K/AKT pathways. Furthermore, 26 different metabolites corresponding to 25 metabolic pathways were identified, and 5 of which were related to the formation of osteoporosis. Interestingly, 23 abnormal metabolites were recovered after 6-AA treatment. ConclusionOur results revealed the significant anti-osteoporotic effects of 6-AA on ovariectomized mice which were probably exerted via suppression of osteoclast formation and bone resorption.

Maternal exposure to cadmium from puberty through lactation induces abnormal reproductive development in female offspring

Four-week-old female ICR mice were exposed to Cd through drinking water from puberty through lactation to investigate the effects of reproductive development in female offspring. Our results showed that maternal Cd exposure from puberty to lactation induced vaginal opening delay, and disturbed estrous cycle in the offspring on postnatal day (PND) 21, without affecting the body weight at vaginal opening. The histopathology results showed the increased primordial follicles and the decreased secondary follicles, and the mRNA level of Amh increased in the offspring’s ovaries upon Cd exposure, suggesting the inhibition of ovarian follicular development on PND21. Moreover, the level of serum estradiol reduced and genes associated with steroidogenesis (3β-Hsd, P450scc and P450arom) were downregulated upon Cd exposure on PND 21. Thus, Cd may inhibit the follicular development via disturbing the mRNA level of genes associated with steroidogenesis and then the synthesis of estradiol in prepuberty. Taken together, despite the lack of attention to estrous cycle at termination, maternal Cd exposure from puberty to lactation induced the adverse effects on reproductive development of female offspring, including the delay of vaginal opening, irregular estrous cycle and inhibition of follicular development, via disturbing the mRNA level of genes associated with follicular development and steroidogenesis.

Acoustic camera system for measuring ultrasound communication in mice

SummaryTo investigate biological mechanisms underlying social behaviors and their deficits, social communication via ultrasonic vocalizations (USVs) in mice has received considerable attention as a powerful experimental model. The advances in sound localization technology have facilitated the analysis of vocal interactions between multiple mice. However, existing sound localization systems are built around distributed-microphone arrays, which require a special recording arena and long processing time. Here, we report a novel acoustic camera system, USVCAM, which enables simpler and faster USV localization and assignment. The system comprises recently developed USV segmentation algorithms with a modification for overlapping vocalizations that results in high accuracy. Using USVCAM, we analyzed USV communications in a conventional home cage, and demonstrated novel vocal interactions in female ICR mice under a resident-intruder paradigm. The extended applicability and usability of USVCAM may facilitate future studies investigating typical and atypical vocal communication and social behaviors, as well as the underlying mechanisms.

Female mouse model of diabetes mellitus induced by streptozotocin and high-carbohydrate high-fat diet.

With the improvement of people's living standards and rapid economic development, the incidence of diabetes mellitus (DM) is increasing in most parts of the world. DM presents an important potential threat to human health. In the present study, a model of diabetes in female mice was established, and fasting blood glucose was detected at week 4, after which the biochemical profiles were evaluated by histopathological analysis. The success rate of modeling in the normal control (NC) group and the low/ middle/high-dose streptozotocin (STZ) group were 0, 0, 25% and 60%, respectively. In the middle-dose and high-dose STZ groups, the liver index was increased significantly compared with the NC group (p⟨0.05). The blood biochemical indicators of total cholesterol and low density lipoprotein cholesterol in three STZ injection groups were as follows: alanine aminotransferase and aspartate transaminase in middle- and high-dose STZ groups, high-density lipoprotein cholesterol and serum creatinine in the high-dose STZ group, and blood urea nitrogen in the middle-dose STZ group were significantly increased (p⟨0.05). The level of total triglycerides was lower, obviously, in the high-dose STZ group than in the NC group (p⟨0.05). The mice showed marked steatosis, green-dyed fiber tissue coloring in varying degrees, and the contour of the hepatic lobules basically disappeared in STZ injection groups. The results suggest that to establish a diabetes model for female ICR mice, the optimum dose of STZ is 100 mg/kg.

SET improved oocyte maturation by serine/threonine protein phosphatase 2A and inhibited oocyte apoptosis in mouse oocytes

SET is a multifunctional protein involved in a variety of molecular processes such as cell apoptosis and cell-cycle regulation. In ovaries SET is predominantly expressed in theca cells and oocytes. In polycystic ovary syndrome (PCOS) patients the expression of SET was increased than healthy people. The current study was designed to determine whether SET plays a role in oocyte maturation and apoptosis, which may provide clues for the underlying pathological mechanism of follicular development in PCOS patients. Oocytes at germinal vesicle (GV) stage were collected from 6-week-old female ICR mice ovaries. The expression of SET was manipulated by AdCMV-SET and AdH1-SiRNA/SET adenoviruses. SET overexpression improved oocyte maturation whereas SET knockdown inhibited oocyte maturation. Moreover, SET negatively regulated serine/threonine protein phosphatase 2A (PP2A) activity in oocytes. Treatment with PP2A inhibitor okadaic acid (OA) promoted oocyte maturation. Furthermore, PP2A knockdown confirmed the role of PP2A in oocyte maturation, and OA was able to block the AdH1-SiRNA/SET-mediated inhibition on oocyte maturation. The central role of PP2A in SET-mediated regulation of oocyte maturation was confirmed by the finding that SET increased the expression of bone morphogenetic protein 15 (BMP15) and growth differentiation factor 9 (GDF9) and PP2A inhibited their expressions. Besides, SET inhibited oocyte apoptosis through decreasing the expression of caspase 3 and caspases 8, while PP2A had no effect on oocyte apoptosis. SET promoted oocyte maturation by inhibiting PP2A activity and inhibited oocyte apoptosis in mouse in-vitro cultured oocytes, which may provide a pathologic pathway leading to impaired oocyte developmental competence in PCOS.

1038-P: Exposures to Hyperglycemia in the Embryonic Period and to Breast Milk from Mothers with Hyperglycemia Affect Glucose Tolerance with Higher Glucagon-Like Peptide-1 Secretion

Exposure to hyperglycemia during pregnancy results in diabetes in offspring. However, its underlying mechanisms are unclear. We therefore focused on the glucose metabolism of the offspring who were exposed to hyperglycemia in uterus or who were breast-fed from mothers with hyperglycemia. ICR female mice received single intraperitoneal injection (i.p.) of streptozotocin at six weeks old as the model of mother with diabetes (MD) . An additional group of mice received citric acid buffer injection, instead, as the model of mother with normal glycemia (MNG) . We established the model exposed to hyperglycemia in the uterus (EHU) by switching offspring from MD to MNG right after birth. As the model exposed to the breast milk of mothers with hyperglycemia (EBM) , the offspring from MNG were moved to MD right after birth. To eliminate the effects of cross fostering, offspring from MNG were fostered by another MNG as the control group. In intraperitoneal glucose tolerance test (IPGTT) at P60, EHU and EBM didn’t show increase of blood glucose level (BG) nor decrease of plasma insulin level but showed in oral glucose tolerance test (OGTT) at P50 (AUC of BG: p &amp;lt; 0.in EHU, p &amp;lt; 0.00in EBM, AUC of plasma insulin level per BG: &amp;lt;0.00in EHU and EBM) . From these conflicting results, we suspected the participation of glucagon-like peptide-1 (GLP-1) . However, the AUC of plasma total GLP-1 and active GLP-1 levels were higher in OGTT. Since we didn’t observe any difference in beta cell area per whole pancreas section by immunostaining, EHU and EBM might have GLP-1 resistance. Disclosure K.Ishii: None. M.Odawara: Other Relationship; Astellas Pharma Inc., AstraZeneca, Daiichi Sankyo, Daiichi Sankyo, Dainippon Pharmaceutical Co., Ltd., Eli Lilly Japan K.K., Kowa Company, Ltd., Kyowa Kirin Co., Ltd., Merck &amp; Co., Inc., Mitsubishi Tanabe Pharma Corporation, Nippon Boehringer Ingelheim Co. Ltd., Novartis Pharma K.K., Novo Nordisk, Omnipod, ONO PHARMACEUTICAL CO., LTD., Sanofi K.K., Taisho Pharmaceutical Holdings Co., Ltd., Takeda Pharmaceutical Co.,Ltd., Teijin Pharma Limited. R.Suzuki: Consultant; Novo Nordisk A/S, Other Relationship; Daiichi Sankyo, Eli Lilly Japan K.K., MSD K.K., Ono Pharmaceutical Co., Ltd., Sanofi K.K., Sumitomo Dainippon Pharma Co., Ltd., Research Support; Boehringer Ingelheim Japan, Inc., LifeScan, Mitsubishi Tanabe Pharma Corporation. J.Sasaki: None. K.Sugai: None. M.Ito: None. H.Iwasaki: None. N.Hara: None. M.Aierken: n/a. G.Li: None. H.Suwanai: None.