BACKGROUND: The therapeutic potential of exosomes from human umbilical cord mesenchymal stem cells (HUMSCsExo) for delivering specific circular RNAs (circRNAs) in treating premature ovarian failure (POF) is not well understood.This study aimed to explore the efficacy of HUMSCs-Exo in delivering hsa_circ_0002021 for POF treatment, focusing on its effects on granulosa cell (GC) senescence and ovarian function. METHODS: Bioinformatic analysis was conducted on circRNA profiles using the GSE97193 dataset from GEO, targeting granulosa cells from varied age groups. To simulate granulosa cell senescence, KGN cells were treated with cyclophosphamide (CTX). HUMSCs were transfected with pcDNA 3.1 vectors to overexpress hsa_circ_0002021, and the HUMSCsExo secreted were isolated. These exosomes were characterized by transmission electron microscopy (TEM) and Western blotting to confirm exosomal markers CD9 and CD63. Co-culture of these exosomes with CTX-treated KGN cells was performed to assess b-galactosidase activity, oxidative stress markers, ROS levels, and apoptosis via flow cytometry.Interaction between hsa_circ_0002021, microRNA-125a-5p (miR-125a-5p), and cyclin-dependent kinase 6 (CDK6) was investigated using dual-luciferase assays and RNA immunoprecipitation (RIP). A POF mouse model was induced with CTX, treated with HUMSCs-Exo, and analyzed histologically and via immunofluorescence staining. Gene expression was quantified using RT-qPCR and Western blot. RESULTS: hsa_circ_0002021 was under expressed in both in vivo and in vitro POF models and was effectively delivered by HUMSCs-Exo to KGN cells, showing a capability to reduce GC senescence. Overexpression of hsa_circ_0002021 in HUMSCs-Exo significantly enhanced these anti-senescence effects. This circRNA acts as a competitive adsorbent of miR-125a-5p, regulating CDK6 expression, which is crucial in modulating cell cycle and apoptosis. Enhanced expression of hsa_circ_0002021 in HUMSCs-Exo ameliorated GC senescence in vitro and improved ovarian function in POF models by modulating oxidative stress and cellular senescence markers. CONCLUSION This study confirms that hsa_circ_0002021, when delivered through HUMSCs-Exo, can significantly mitigate GC senescence and restore ovarian function in POF models. These findings provide new insights into the molecular mechanisms of POF and highlight the therapeutic potential of circRNA-enriched exosomes in treating ovarian aging and dysfunction.

The latest epidemiological data suggests that the situation of adult diabetes in China is severe, and metabolic diseases have become significant chronic illnesses that have a serious impact on public health and social development. After more than six years of practice, the National Metabolic Management Center(MMC) has developed distinctive approaches to manage metabolic patients and has achieved a series of positive outcomes, continuously advancing the standardized diagnosis and treatment model. In order to further improve the efficiency, based on the first edition, the second edition guideline was composed by incorporating experience of the past six years in conjunction with the latest international and domestic guidelines.

It is known that low-frequency pulsed electromagnetic fields (PEMFs) can promote the differentiation and maturation of rat calvarial osteoblasts (ROBs) cultured in vitro. However, the mechanism that how ROBs perceive the physical signals of PEMFs and initiate osteogenic differentiation remains unknown. In this study, we investigated the relationship between the promotion of osteogenic differentiation of ROBs by 0.6 mT 50 Hz PEMFs and the presence of polycystin2 (PC2) located on the primary cilia on the surface of ROBs. First, immunofluorescence staining was used to study whether PC2 is located in the primary cilia of ROBs, and then the changes of PC2 protein expression in ROBs upon treatment with PEMFs for different time were detected by Western blotting. Subsequently, we detected the expression of PC2 protein by Western blotting and the effect of PEMFs on the activity of alkaline phosphatase (ALP), as well as the expression of Runx-2, Bmp-2, Col-1 and Osx proteins and genes related to bone formation after pretreating ROBs with amiloride HCl (AMI), a PC2 blocker. Moreover, we detected the expression of genes related to bone formation after inhibiting the expression of PC2 in ROBs using RNA interference. The results showed that PC2 was localized on the primary cilia of ROBs, and PEMFs treatment increased the expression of PC2 protein. When PC2 was blocked by AMI, PEMFs could no longer increase PC2 protein expression and ALP activity, and the promotion effect of PEMFs on osteogenic related protein and gene expression was also offset. After inhibiting the expression of PC2 using RNA interference, PEMFs can no longer increase the expression of genes related to bone formation. The results showed that PC2, located on the surface of primary cilia of osteoblasts, plays an indispensable role in perceiving and transmitting the physical signals from PEMFs, and the promotion of osteogenic differentiation of ROBs by PEMFs depends on the existence of PC2. This study may help to elucidate the mechanism underlying the promotion of bone formation and osteoporosis treatment in low-frequency PEMFs.
Alkaline Phosphatase/metabolism* ; Animals ; Electromagnetic Fields ; Osteoblasts/metabolism* ; Osteogenesis/genetics* ; Rats ; TRPP Cation Channels/physiology*

Objective:To investigate the role of neurite outgrowth inhibitor (Nogo)-oligodendrocyte myelin glycoprotein (Omgp)/Ras homologous (Rho)-Rho-associated coiled-coil forming protein kinase (Rock) signaling pathway in acute brain injury of carbon monoxide (CO) poisoning rats and treatment feasibility with Rho kinase inhibitor hydrochloride fasudil.Methods:According to random number table method, 135 healthy male SD rats were divided into three groups: a normal control group, a CO poisoning group and a fasudil treatment group ( n=45). Rat models of acute severe CO poisoning were established in the CO poisoning group and fasudil treatment group by inhalation method in a hyperbaric oxygen chamber. All rats received hyperbaric oxygen therapy for two weeks. Rats in the farsudil treatment group were intraperitoneally injected with hydrochloride farsudil for intervention (15 mg/[kg·d], once a d for 2 weeks), while those in the CO poisoning and normal control groups received the same volume of normal saline. The ultrastructures of rat brain tissues were observed by transmission electron microscopy one week after modeling. Staining intensities of Nogo- and OMgp-positive cells were detected by immunohistochemistry, and those of Rock-positive cells were analyzed by immunofluorescence one d, one week, one month and two months after modeling. The protein expressions of Nogo, OMgp and Rock in brain tissues were detected by Western blotting one d, one week, one month and two months after modeling. Results:In the CO poisoning group, the ultrastructures of brain tissues and blood-brain barrier were damaged obviously, and the changes in nucleus, mitochondria and synaptic structure were obvious; while fasudil treatment could effectively maintain the integrity of ultrastructures and functions of brain tissues, and reduce brain edema. One d, one week, one month and two months after modeling, the staining intensities of Nogo, OMgp and Rock positive cells and protein expression levels of Nogo, OMgp and Rock in the CO poisoning group were significantly higher than those in the normal control group at the same time point ( P<0.05); the staining intensities of Nogo, OMgp and Rock positive cells and protein expression levels of Nogo, OMgp and Rock in the fasudil treatment group were significantly lower than those in the CO poisoning group at the same time point ( P<0.05). Conclusion:The activation of Nogo-OMgp/Rho-Rock signaling pathway related molecules (Nogo, OMgp and Rock) is closely related to acute brain injury caused by CO poisoning; hydrochloride fasudil can effectively down-regulate the protein expressions of Nogo, OMgp and Rock, therefore obviously alleviate brain injury after CO poisoning.

Objective To investigate the therapeutic effects of Xingzhi Yinao (XZYN) particles combined with hyperbaric oxygen therapy on cognitive impairment and motor dysfunction in patients with delayed encephalopathy after acute carbon monoxide poisoning (DEACMP). Methods Sixty-seven patients with DEACMP were admitted to the Affiliated Yantai Yuhuangding Hospital of Qingdao University from January 2011 to December 2015, and they were randomly divided into a control group (given conventional treatment such as inhalation of oxygen, cytidine diphosphate cholin and vitamin B, 19 cases), a hyperbaric oxygen (HBO) treatment group (given conventional treatment + hyperbaric oxygen therapy once a day, 24 cases) and a XZYN particles treatment (XZYN group, given conventional treatment, hyperbaric oxygen and XZYN particles, 24 cases), the therapeutic course being 2 months in the three groups. Before and after treatment for 1 and 2 months, the cognitive function and motor function of the patients were evaluated by the use of activity of daily living (ADL) scale, Montreal cognitive assessment (MoCA) scale, and mini-mental state examination (MMSE) scale; the severity of cerebral white matter injury was assessed by age related white matter changes (ARWMC) scale; and the electromyographic evoked potential was used to detect the amplitude and latency of P300 to assess the severity of cognition impairment and prognosis. Results With the prolongation of therapeutic time, after treatment, the neurological function scores of ADL, MoCA, MMSE and amplitude of P300 were increased, while ARWMC was decreased and the latency of P300 was shortened gradually in the three groups, and the changes of above indexes after treatment for 2 months in XZYN group were more significant than those in either HBO group or control group[ADL score: 70.2±8.3 vs. 60.5±8.1, 23.0±6.1, MoCA score: 26.1±3.1 vs. 22.2±2.7, 18.2±3.6, MMSE score:25.9±4.1 vs. 22.4±3.5, 18.1±4.5, ARWMC score: 7.0±2.1 vs. 8.7±2.2, 15.2±3.3, latency of P300 (ms):332.9±20.4 vs. 352.5±23.6, 381.7±30.3, amplitude of P300 (μV): 6.5±1.6 vs. 5.6±1.3, 4.1±1.5, all P < 0.05]. Conclusion The hyperbaric oxygen therapy combined with XZYN particles for treatment of patients with DEACMP can significantly improve their cognitive and motor functions and ameliorate the severity of cerebral white matter injury.

Objective To explore the effects of N-butylphthalide on the expressions of ZO-1 and claudin-5 in blood-brain barrier (BBB) in rats with acute carbon monoxide (CO) poisoning. Methods A total of 144 adult healthy male Sprague-Dawley (SD) rats were randomly divided into normal control group, CO poisoning group, and NBP treatment group, with 48 rats in each group. The acute CO poisoning model was reproduced in hyperbaric oxygen chamber, and all model rats were given hyperbaric oxygen therapy once daily. The rats in the normal control group were free to breathe fresh air. The rats in NBP treatment group were administered orally NBP 60 mg/kg twice a day at 2 hours after poisoning until death. The rats in normal control group and CO poisoning group were treated with equal amount of pure olive oil. Four rats were sacrificed from each group at 1, 3, 7, 14 days after model reproducing, respectively. The changes in ultrastructure of BBB were observed under transmission electron microscope. The expressions of ZO-1 and claudin-5 proteins were determined by immunofluorescence staining and Western Blot. The localization of the two target proteins was observed by immunofluorescence double staining. The correlation between the two proteins was analyzed by linear regression. Results The ultrastructure of BBB was normal in normal control group, some ZO-1 and a large number of claudin-5 positive cells were observed. The ultrastructure of BBB was seriously injured, ZO-1 and claudin-5 positive cells in brain tissue were significantly decreased, and the expressions of ZO-1 and claudin-5 proteins in brain tissue at 1 day after poisoning in CO poisoning group were significantly lower than those of normal control group (ZO-1 protein:3.38±0.30 vs. 24.50±5.62, claudin-5 protein: 11.38±0.93 vs. 46.35±6.88, both P < 0.05), and although gradually restored, they were maintained at relatively lower levels until 14 days as compared with those in normal control group (ZO-1 protein: 10.35±0.80 vs. 24.63±3.57, claudin-5 protein: 32.35±3.11 vs. 46.43±7.20, both P < 0.05). NBP treatment could significantly alleviate the ultrastructure injury of BBB induced by acute CO poisoning, the amount of ZO-1 and claudin-5 positive cells in brain tissue were significantly increased, as well as the expressions of ZO-1 and claudin-5 proteins were significantly increased, which were significantly higher than those of CO poisoning group from 1 day and 3 days on, respectively (1-day ZO-1 protein: 7.57±0.69 vs. 3.38±0.30, 3-day claudin-5 protein:20.46±1.42 vs. 11.43±0.86, both P < 0.05), and which showed an increase tendency with time prolongation. The results of immunofluorescence double staining showed that ZO-1 and claudin-5 proteins could not only coexist in the same cell, but also could be expressed separately in different cells. Linear regression analysis showed the positive correlation between the expressions of ZO-1 and claudin-5 proteins in brain tissue of rats with acute CO poisoning (R2= 0.917, P = 0.022). Conclusion NBP could markedly improve the ultrastructure and functional integrity of BBB through up-regulating the expressions of ZO-1 and claudin-5 proteins, and then reduce brain damage caused by CO poisoning.

Objective To explore the differential expression of lncRNAs in patients with depression and its relationship with personality traits and social support.Methods The differential expression of lncRNAs in 5 patients with depression (MDD) and 5 normal controls (NC) was screened by gene chip.To validate the gene chip dataset,qRT-PCR was used to verify the expression levels of these 10 lncRNAs in a separate set of 138 consecutive patients and 43 normal subjects,and then the relationships of lncRNA expression level with personality traits and social support were analyzed.Results A total of 2 649 lncRNAs were differentially expressed,of which 534 were up-regulated and 2 115 down-regulated.The expression levels of 8 lncRNAs analyzed by qRT-PCR in patients with depression were significantly down-regulated (P＜0.05 or P＜0.01).The △Ct value of PY4 was negatively correlated with anxiety/somatization factor (r=-0.210,P＜0.05),and the △Ct values of PY1,PY2 and PY6 were negatively correlated with the social support availability (r=-0.383,-0.391,-0.381 all P＜0.05).Apart from PY1,the △Ct values of the other lncRNAs were negatively correlated with paranoid (P＜0.05),the △Ct values of PY3,PY6 and PY9 were negatively correlated with the borderline and obsessive-compulsive (P＜0.05) and except for PY10,the △Ct values of the other lncRNAs were negatively correlated with the schizoid (P＜0.05).Conclusion The expression levels of theses 8 lncRNAs are significantly down-regulated in patients with depression,and there is a certain correlation with anxiety/somatization factor,personality traits and social support.

Objective To explore the neuroprotective effect of targeted regulation Nrf2 gene on rats with brain injury caused by acute severe carbon monoxide ( CO ) poisoning. Methods A total of 180 healthy adult SD rats were divided into 4 groups at random:normal control group( NC group) ,CO poisoning group(CO group),lentivirus group(LV group) and Nrf2 gene therapy group(Nrf2 group),and 45 rats in each group. An acute CO toxic rat model was established by inhalation in a hyperbaric oxygen tank. The lentivirus group was directly injected with lentivirus dilution (4×106 TU/μl) into striatum with a microsy-ringe guided by a stereotactic apparatus,and the Nrf2 gene therapy group was administrated the same dose of recombinant Nrf2 gene lentivirus dilution,while rats in the normal control group and the CO poisoning group were received the same amount of normal saline. Five rats were taken and decapitated at day 1,day 7 and week 2 from each group,respectively. The mitochondrial membrane potential (MMP) of neurons in brain tis-sue was detected by JC-1 method,and the expressions of Nrf2 and GCLC proteins were observed by immuno-histochemistry and Western Blot. Results Compared with the NC group (cortex:(75. 3±6. 8);hippocam-pus:(76. 4±7. 1);striatum:(73. 8±7. 3)) at the same time point,the MMPs of neurons in CO group (cor-tex:(34. 5±6. 7);hippocampus:(30. 3±5. 6);striatum:(41. 5±6. 1) and LV group (cortex:(36. 8±6. 2);hippocampus:(30. 8±6. 0);striatum:(42. 7±6. 3)) were significantly decreased,and the difference was sig-nificant(P<0. 05). However,there was no significant difference between the CO poisoning group and the lentivirus group (P>0. 05). A small amount of Nrf2 protein (0. 22±0. 05) and GCLC protein (0. 24±0. 04) were expressed in the brain tissue of normal control rats. The expressions of Nrf2 protein (0. 31±0. 06,0. 31 ±0. 05) and GCLC protein (0. 30±0. 04,0. 31±0. 07) in CO group and LV group were slightly increased (P<0. 05). Similarly,there was no significant difference between the CO poisoning group and the lentivirus group (P>0. 05). The MMPs value of nerve cells in the Nrf2 group (cortex:(53. 3±5. 3);hippocampus:(56. 9±6. 1);striatum:(60. 6±6. 0)) also decreased,but it was significantly higher than that in the CO group and the LV group at the same time point (P<0. 05) . The expression of Nrf2 in brain tissue was signifi-cantly increased (0. 59±0. 05),and there was significant difference between CO group and LV group at the same time point (P<0. 05);GCLC protein increased slightly (0. 37±0. 06),but there was no statistical difference compared with CO poisoning group and lentivirus group (P>0. 05). Conclusion CO poisoning could induce oxidative stress and damage mitochondrial function of nerve cells. The active state of targeted regulation Nrf2 could significantly enhance the antioxidant capacity of rats and positively protect rats against brain injury induced by acute severe CO poisoning.

Objective Atherosclerosis ( AS) is a common pathological basis of cardiovascular diseases.Adiponec-tin ( APN) has been shown to have an anti-AS effect, and the underlying mechanisms, however, are largely unknown.Nu-clear transcription factorκB ( NF-κB) has also been regarded as a proatherogenic factor, mainly because of its regulation of a variety of the proinflammatory genes linked to AS.It is hypothesized that the inhibitory effects of APN on AS is through the inhibition of NF-κB signaling pathway.The aim of this study was to test the hypothesis via investigation and validation of the inhibitory effect of APN on AS in ApoE-/-mice, and to delineate the roles of NF-κB signaling pathway in modulating the APN effect on AS in vivo.Methods APN overexpression in ApoE-/-mice were mediated by transfecting adenovirus bearing a vector encoding for APN and enhanced green fluorescent protein ( Ad-APN-eGFP) .The AS in ApoE-/-mice was induced by feeding a high-fat diet.To validate the inhibitory effect of the adenovirus mediated APN overexpression on AS in the ApoE-/-mice.120 male ApoE-/-mice aged 12 weeks were randomly and evenly assigned into two groups (60 mice per group), and were fed with a high-fat diet to induce AS.At 0 day, 2, 4, and 6 weeks of high-fat diet feeding.The 2 groups of mice were injected intravenously in the tail with either 100 μL (3.0 ×108 p.f.u) of Ad-eGFP virus ( control group) or the same amount of Ad-APN-eGFP virus ( APN group) .Blood samples and aortic tissues were taken at 0 day, 4, and 8 weeks of high-fat diet feeding.For the blood samples, FABA was used to analyze the concentrations of blood lipids and ELIZA was used to test the concentrations of serum APN.For the aortic tissues, oil red O staining was used to detect the surface lesion percentage.Masson staining was used to evaluate the collagen content and fibrous cap thickness of the plaque area.Immunofluorescence method was used to detect APN and NF-κB p65 expression.Western blot was used to de-tect the expressions of APN,nuclear NF-κB p65 and the downstream factors of NF-κB pathway.Results APN inhibited the formation of atherosclerotic plaque in ApoE-/-mice.The lesion formation in aortic sinus was significantly inhibited ( P<0.01).Compared with the control group, the oil red O staining showed that the surface area ratio of atherosclerotic le-sions was decreased significantly in the Ad-APN group ( P<0.001 ): the percentage of surface lesions in the 4 weeks groups was 27.78 ±8.64 vs.33.02 ±5.18 (%);the 8 weeks groups was 31.58 ±5.87 vs.52.16 ±5.79 (%) .As the serum APN was increased,the concentration of TC, TG and LDL-C were significantly decreased( P<0.001 for all) , and the growth of body weight was slowed down(P<0.05).APN effectively inhibited the expression of NF-κB nuclear protein p65 and inflammatory factors.Conclusions Adiponectin reduces the inflammatory reactions in atherosclerosis through in-hibiting the NF-κB pathway.

OBJECTIVETo explore whether miR-200b suppresses tumor cell invasion by targeting PROM1, thus to reveal the molecular mechanism that miR-200b functions as a tumor suppressor in glioma.

METHODSPROM1 3'UTR-luciferase vector was constructed and dual-luciferase reporter gene assay was employed to examine the effect of miR-200b on luciferase activity. Human glioblastoma U87 cells were transfected with miR-200b mimics, and next qRT-PCR and Western blotting were performed to detect the expressions of PROM1 mRNA and protein. The effect of PROM1 down-regulation on invasion was observed after PROM1 siRNA were transfected into U87 cells.

RESULTSThe miR-200b bound to the 3'-untranslated region (UTR) of PROM1 and inhibited the luciferase activity. Its luciferase activity was down-regulated by 57.0% (P < 0.01). PROM1 protein and mRNA expressions were significantly down-regulated when miR-200b was overexpressed in the U87 cells (P < 0.05). siRNA-mediated down-regulation of PROM1 suppressed the potential of cell invasion. The invasion ability of SKOV3 cells after transfection with siRNA-PROM1 was significantly lower than that in the negative control cells (P < 0.05).

CONCLUSIONmiR-200b may suppress cell invasion by targeting PROM1 in glioma.

3' Untranslated Regions ; AC133 Antigen ; Antigens, CD ; metabolism ; Cell Line, Tumor ; Down-Regulation ; Genes, Reporter ; Genes, Tumor Suppressor ; Genetic Vectors ; Glioblastoma ; genetics ; metabolism ; Glycoproteins ; metabolism ; Humans ; Luciferases ; MicroRNAs ; metabolism ; Peptides ; metabolism ; RNA, Messenger ; RNA, Small Interfering ; Transfection