Identification of Circular RNAs in Hypothalamus of Gilts during the Onset of Puberty.

The disorders of puberty have shown negative outcomes on health of mammals, and the hypothalamus is thought to be the main regulator of puberty by releasing GnRH. Many studies show that the circular RNAs (circRNAs) might be implicated in the timing of puberty in mammals. However, the circRNAs in the hypothalamus of gilts have not been explored. To profile the changes and biological functions of circRNAs in the hypothalamus during the onset of puberty, RNA-seq was utilized to establish pre-, in-, and post-pubertal hypothalamic circRNAs profiles. In this study, the functions of hypothalamic circRNAs were enriched in the signaling pathway of neurotrophin, progesterone-mediated oocyte maturation, oocyte meiosis, insulin, ErbB, and mTOR, which have been highly suggested to be involved in the timing of puberty. Furthermore, 53 circRNAs were identified to be putative hypothalamus-specific expressed circRNAs, and some of them were exclusively expressed in the one of three pubertal stages. Moreover, 22 differentially expressed circRNAs were identified and chosen to construct the circRNA-miRNA-gene network. Moreover, 10 circRNAs were found to be driven by six puberty-related genes (ESR1, NF1, APP, ENPP2, ARNT, and DICER1). Subsequently, the expression changes of several circRNAs were confirmed by RT-qPCR. Collectively, the preliminary results of hypothalamic circRNAs provided useful information for the investigation of the molecular mechanism for the timing of puberty in gilts.

Label-free and high-throughput bioluminescence detection of uracil-DNA glycosylase in cancer cells through tricyclic cascade signal amplification.

We develop a label-free and high-throughput bioluminescence method for the sensitive detection of uracil DNA glycosylase (UDG) through enzyme-mediated tricyclic cascade signal amplification. This method exhibits high sensitivity with a detection limit as low as 0.00031 U mL-1, and it can be further applied for the measurement of enzyme kinetic parameters and the screening of UDG inhibitors as well as cancer cell analysis.

[Optimize aqueous extract protocol for Yugubao formula by orthogonal experiment design].

To determine the optimum aqueous extract protocol for Yugubao traditional Chinese medicines formula by using orthogonal experiment design. Through serum pharmacology research, L9(34) orthogonal design with single factor investigation was used to optimize the aqueous extract protocol for Yugubao formula. The effect of water extraction on activity of alkaline phosphatase (ALP) in osteoblast was referred as the evaluation index for investigating four factors: water consumption (A), heating time (B), soaking time (C), and number of decocting (D), analyzing the optimum extraction conditions, and verifying the effectiveness of this process. The optimum aqueous extract protocol for Yugubao was as follows: adding 8 times water into Chinese medical materials, heating for 60 min, soaking for 30 min, and decocting for 1 time. The drug serum of this aqueous extract of Yugubao could significantly up-regulate the osteogenic genes expression. The optimum aqueous extract protocol for Yugubao formula was established in this experiment, providing evidence for the development and utilization of Yugubao traditional Chinese medicines formula.

Effects of unfractionated heparin on renal osteodystrophy and vascular calcification in chronic kidney disease rats.

Unfractionated heparin (UFH) is the most widely used anticoagulant in hemodialysis for chronic kidney disease (CKD) patients. Many studies have verified that UFH can induce bone loss in subjects with normal bone, but few have focused on its effect on renal osteodystrophy. We therefore investigated this issue in adenine-induced CKD rats. As CKD also impairs mineral metabolism systemically, we also studied the impacts of UFH on serum markers of CKD-mineral and bone disorder (CKD-MBD) and vascular calcification. We administered low and high doses of UFH (1U/g and 2U/g body weight, respectively) to CKD rats and compared them with CKD controls. At sacrifice, the serum markers of CKD-MBD did not significantly differ among the two UFH CKD groups and the CKD control group. The mean bone mineral densities (BMDs) of the total femur and a region of interest (ROI) constituted of trabecular and cortical bone were lower in the high-dose UFH (H-UFH) CKD group than in the CKD control group (P<0.05 and P<0.01, respectively). The BMD of the femoral ROI constituted of cortical bone did not differ between the H-UFH CKD group and the CKD control group. Histomorphometrical changes in the CKD rats indicated secondary hyperparathyroidism, and the femoral trabecular bone volume, but not cortical bone volume, significantly decreased with increasing UFH dose. The same decreasing trend was found in osteoblast parameters, and an increasing trend was found in osteoclast parameters; however, most differences were not significant. Moreover, no distinct statistical differences were found in the comparison of vascular calcium or phosphorus content among the CKD control group and the two UFH CKD groups. Therefore, we concluded that UFH could induce bone loss in CKD rats with secondary hyperparathyroidism, mainly by reducing the trabecular volume and had little effect on cortical bone volume. The underlying mechanism might involve inhibition of osteoblast activity and promotion of osteoclast activity by UFH. We did not find any effect of UFH on vascular calcification in CKD rats with secondary hyperparathyroidism.

[Effects of drug serum of plantaginis semen on bone formation function of rat osteoblast in vitro].

OBJECTIVE: To study the influence of Plantaginis Semen on cell proliferation, differentiation and function of rat osteoblasts, and investigate the regulation effects of rat osteoblast epithelial sodium channel (ENaC) on bone formation. METHODS: The animal serum was prepared by serum pharmacology means. The cells were got by separating and inducing the SD neonatal rat's skull bone. Cell proliferation and differentiation were evaluated by CCK-8 assay kit and AKP assay kit respectively. Regulation effects on mRNA expression of ENaC and osteogenesis gene were investigated by semi-quantitative PCR. RESULTS: Plantaginis Semen stimulated the osteoblasts proliferation and differentiation,the difference between treatment group and control group had statistical significance (P < 0.01) in a dose-dependent manner. The effects of Plantaginis Semen serum on alpha-ENaC gene expression paralleled those on osteogenic gene (OC, ALP, OP) expression level. CONCLUSION: Plantaginis Semen stimulates proliferation, differentiation and the mRNA expression of ENaC and osteogenesis gene in rat osteoblasts. Our results suggest that ENaC participate in the effects of Plantaginis Semen serum on osteoblast bone formation. Regulation of ENaC channel expression and function may provide a new clue for research on treatment of osteoporosis with traditional Chinese medicine.

[Community structure and biological integrity of periphyton in Hunhe River water system of Liaoning Province, Northeast China].

Taking the Hunhe River water system in Liaoning Province of Northeast China as a case, this paper investigated the community structure of periphyton and the physical and chemical characteristics of water environment at 62 sampling sites, and, by using the biological integrity index of periphyton (P-IBI) and the habitat environment quality index (QHEI), a health assessment on the aquatic ecosystem of the water system was conducted. There was a great spatial heterogeneity in the periphyton community structure, and the sampling sites could be divided into 4 groups, with significant differences in the community structure, species richness, density, and Shannon diversity index among the groups. Canonical correspondence analysis (CCA) showed that ammonia nitrogen (NH4(+)-N) and dissolved phosphorus (PO4(3-)-P) were the main water environment factors driving the formation of the periphyton community structure in the water system. The health assessment of the water system based on P-IBI and QHEI was basically in consistency, though the assessment of several reaches had large discrepancy. Overall, the Suzi River at the upper reaches of the water system had a better health status, while the upper reaches of Hunhe River and the main stream of Hunhe River at its middle and lower reaches were worse in health.

[Effects and mechanism of drug serum of prepared radix polygoni multiflori on rat osteoblast in vitro].

OBJECTIVE: To study the effect of drug serum of prepared Radix Polygoni Multiflori on rat osteoblast (Ob) and its mechanism. METHODS: The animal serum was prepared by serum pharmacology means. The cells were getting by separating and inducing the SD neonatal rat skull bone. The proliferation and differentiation of Ob were detected by CCK-8, alkaline phosphatase (ALP) activity analysis. And RT-PCR method was used to determine the osteogenesis-related genes expression. RESULTS: Compared with control group, the groups with drug serum of prepared Radix Polygoni Mutiflori, 10%, 20% and 30% had an effect on promotion the proliferation significantly on Ob (P < 0.01). There was no concentration-related manner among groups. The 5% and 10% drug serum decreased ALP activity at the post-translation phase compared with control group, but higher doses (20% and 30%) did not have the same effect. However, drug serum of PR/MIN increased significantly osteogenesis-related genes (OC, ALP, Cbfalpha1) mRNA expression (P < 0.05). CONCLUSION: The drug serum of prepared Radix Polygoni Multiflori can stimulate osteoblast proliferation in vitro, and its mechanism may be associated with increasing osteogenesis-related genes expression.

[Study on extraction process for the icariin of hugu capsule with ultrasonic technology].

OBJECTIVE: To study the best ultrasonic technology of the extraction of icariin of Hugu capsule. METHODS: Used the content of icariin as index, orthogonal experiment was carried out to investigate 4 influential factors as follows: the ultrasonic power (A), the ultrasonic frequency (B), the material fluid ratio (C), the time (D). RESULTS: The best extraction conditions were as follows: the ultrasonic power was 120 W, the ultrasonic frequency was 28 KHz, solid-liquid ratio of 1 : 35, the extraction time was 10 min. CONCLUSION: Optimization of extracting process is simple, quick and low energy consumption. Under these conditions, the extraction of icariin is 1.8 times higher than that of the traditional extraction method.

[Piperazinyl estrone prevents bone loss in ovariectomized rats].

AIM: To determine the effect of piperazinyl estrone, a new estrogen derivative, on bone turnover, bone mass and uteri in ovariectomized rats. METHODS: Female Sprague-Dawley rats were ovariectomized (OVX) or sham operated (sham) at the age of 3 months and treated with estrone (E) at 0.75 mg.kg-1.d-1, or with piperazinyl estrone (P-E) at 1 or 10 mg.kg-1.d-1, orally, for 3 months. At the time of death, the uterine weight was measured. Bone histomorphometric analysis of proximal tibial metaphyses (PTM) was performed in undecalcified sections. RESULTS: Bone histomorphometric data showed that the percent trabecular area (% Tb.Ar) of OVX rats with bone high turnover was significantly decreased. The uteri were atrophied. The percent trabecular area (% Tb.Ar) of estrone treated group was increased in decreasing bone turnover manner. But the size and weight of uteri in this group were increased vs OVX group. The bone loss induced by OVX was preserved by P-E treatment, but the mechanism of maintaining bone is different from that of E-treated rats. P-E treatment in low dose did not decrease any bone formation indices, such as percent labeling perimeter, bone formation rate per bone volume (BFR/BV), except bone mineral apposition rate (MAR) compared with E-treated group, and maintained them at OVX level. The uteri were found to be in atrophy compared with the match dose (0.75 mg) of E-treated OVX rats. But rats treated with high dose of P-E showed the same change like E-treated group. CONCLUSION: The finding of this study shows that lower dosage of piperazinyl estrone has effect on preventing the bone losses in OVX rats, while the bone formation and the uterus are not affected, thus supporting the hypothesis that piperazinyl estrone has the potential to prevent postmenopausal bone loss in women with less side effects.

[Bone mineral density and biomechanical effects of Chinese herbal drug Yanhuogubao on experimental osteoporosis in rats].

OBJECTIVE: To explore bone mineral density and biomechanical effects of Chinese herbal drug Yanhuogubao on steroid hormone-induced osteoporosis in rats. METHODS: Male, 3-month-old Sprague-Dawley rats were randomly divided into 4 groups (n = 7), the basal group, the age control group, the hormone group (4.5 mg/kg prednisone acetate, twice a week) and Yanhuogubao (1.5 g/kg/d) group. The bone mineral densities and the mechanical characters measurements of femurs and 5th lumbar vertebrae in every group were made by using DEXA and SWD-10 electronic universal material testing machine. RESULTS: The bone mineral densities on femora and 5th lumbar vertebrae in hormone group decreased, and the loads imposed on the femoral trunk in three-point bending test decreased by 17.1% (P < 0.05), though the rest parameters tended to decrease in comparison with control group. In the meantime, Yanhuogubao can prevent prednisone acetate-induced the bone mineral densities and mechanical characters of femurs and lumbar vertebrae in rats from decline, compared with hormone group. CONCLUSION: When steroid hormone was used for a long time, the bone mineral densities and mechanical characters of femora and 5th lumbar vertebrae in the rats would be led to decrease, on the contrary, Yanhuogubao could arrest the decline of bone mineral densities and mechanical characters of rat skeleton.

[Combination of ginsenosides with low dose estrogen showed synergetic effect on ovariectomy induced osteopenia in rats].

AIM: To determine whether low dose of estrogen in combination with ginsenosides can completely prevent bone loss in ovariectomized rats. METHODS: Four-month-old ovariectomized rats were treated either with 100 and 300 mg.kg-1 of ginsenosides or 30 and 100 micrograms.kg-1 of 17 alpha-ethynylestradiol alone, or ginsenosides 100 mg.kg-1 in combination with 17 alpha-ethynylestradiol 30 micrograms.kg-1 for 10 weeks. Double in vivo fluorochrome labeling was made. The undecalcified longitudinal proximal tibial metaphyseal sections were processed and stained with Goldner's trichrome for histomorphometric analysis of the bone. RESULTS: Body weights and serum cholesterol were increased in ovariectomized (OVX) rats. The rats lost 74% of bone volume and high bone turnover was induced after OVX compared with the sham group. Bone volume increased by 205% in the high dose estrogen treated group while it was increased by 105% in the low dose group. The two doses of estrogen were shown to inhibit osteoclasts surface (by -65% and -55%, P < 0.01) and decrease bone turnover rate (by -85% and -83%, P < 0.01). High dose of estrogen was found to inhibit growth and stimulate uterine weight gain in rats while low dose did not. High dose of ginsenosides increased bone volume by 84% (P < 0.01) and decreased bone turnover rate by -64% (P < 0.05) while lower dose of ginsenosides did not. However, low dose ginsenosides combined with low dose estrogen achieved well preventive effects: increase of 202% in bone volume, decrease of 66% in bone turnover rate and 72% in osteoclasts surface. The combined effect in preventing bone loss equals to that the high dose of estrogen alone did. CONCLUSION: Use of low dose of estrogen plus ginsenosides showed synergistic effect on prevention of osteoporosis induced by ovariectomy.