Influence of miR-155 on behaviors of depression mice through regulating Wnt/ß-catenin signaling pathway.

OBJECTIVE: To study the influence of micro ribonucleic acid (miR)-155 on depression-like behaviors of depression mice, and to explore the role of Wnt/b-catenin signaling pathway in behavioral regulation of depression mice. MATERIALS AND METHODS: The mouse model of depression was established via chronic unpredictable mild stress (CUMS). All mice were randomly divided into control group (n=12), model group (n=12), and fluoxetine group (n=12). The expression level of miR-155 in the hippocampus of mice in each group was detected via quantitative Polymerase Chain Reaction (qPCR). The changes in the behaviors of mice in each group were evaluated via behavioral experiments. The apoptosis level in the hippocampus of mice in each group was detected via terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining. Moreover, the content of inflammatory factors in the hippocampus of mice in each group was detected using the enzyme-linked immunosorbent assay (ELISA) kits. The expression levels of Wnt/b-catenin signaling pathway-related proteins in each group were detected via Western blotting. RESULTS: The expression level of miR-155 in the hippocampus was significantly higher in model group than that in control group (p<0.01). Meanwhile, the expression level of miR-155 was significantly lower in fluoxetine group than that in model group (p<0.01). There were no statistically significant differences in the crossing score and rearing score in the open field test among groups (p>0.05). Compared with those in control group, the immobility time in tail suspension test and forced swimming test were significantly increased (p<0.01), while the sucrose preference degree significantly declined (p<0.01) in model group. Fluoxetine could significantly reduce the immobility time in tail suspension test and forced swimming test (p<0.01) and increase the sucrose preference degree (p<0.01) in model group. The number of TUNEL-positive cells in the hippocampus of mice in model group was significantly larger than that in control group (p<0.01). Fluoxetine could effectively reduce the number of TUNEL-positive cells in the hippocampus (p<0.01). Compared with those in control group, the content of tumor necrosis factor-α (TNF-a), interleukin-1b (IL-1b), and IL-6 in the hippocampus was significantly increased (p<0.01), while the content of IL-10 was significantly decreased (p<0.01) in model group. Fluoxetine could effectively reduce the content of TNF-a, IL-1b, and IL-6 (p<0.01) and increase the content of IL-10 (p<0.01). Besides, in model group, the expression levels of dishevelled-1 (DVL-1) and b-catenin in hippocampus remarkably declined (p<0.01), while the expression levels of glycogen synthase kinase-3b (GSK-3b) and adenomatous polyposis coli (APC) were remarkably increased (p<0.01) compared with those in control group. Fluoxetine could effectively lower the expressions of GSK-3b and APC in the hippocampus (p<0.01) and increase the expressions of DVL-1 and b-catenin (p<0.01) in model group. CONCLUSIONS: MiR-155 is involved in regulating the depression-like behaviors of depression mice through promoting the release of inflammatory factors and the apoptosis of hippocampal neurons. Its mechanism may be related to the inhibition of the Wnt/b-catenin signaling pathway.

Effects of lncRNA BANCR on endometriosis through ERK/MAPK pathway.

OBJECTIVE: To investigate the regulatory role of long non-coding ribonucleic acid (lncRNA) BRAF-activated non-coding RNA (BANCR) in rats with endometriosis (EMs) and its mechanism of action. MATERIALS AND METHODS: A total of 30 healthy, unmated, female Sprague-Dawley (SD) rats were selected and divided into sham-operation group, model group and lncRNA BANCR intervention group, and a rat model of EMs was established by means of autotransplantation. The volume of eutopic endometrium in each group of rats was measured, and hematoxylin and eosin (HE) staining was applied to detect the impacts on the pathological morphology of ectopic endometrial tissues in each group. The levels of vascular endothelial growth factor (VEGF), matrix metalloproteinase-2 (MMP-2) and MMP-9 in the rat serum were determined by virtue of enzyme-linked immunosorbent assay (ELISA), reverse transcription-polymerase chain reaction (RT-PCR) was performed to measure the messenger RNA (mRNA) levels of extracellular signal-regulated kinase (ERK) and mitogen-activated protein kinase (MAPK) in the uterine tissues in each group of rats, and Western blotting assay was adopted to detect the levels of phosphorylated ERK and MAPK proteins in the rat uterine tissues in each group. RESULTS: Compared with those in sham-operation group, the volume of eutopic endometrium in the rats was increased markedly, the pathological morphology was poorer, and the content of VEGF, MMP-2 and MMP-9 in the serum, the mRNA levels of ERK and MAPK in the uterine tissues, and the levels of phosphorylated ERK and MAPK proteins were elevated notably in model group. The rats in lncRNA BANCR intervention group had evidently decreased volume of eutopic endometrium, improved pathological morphology and significantly declined content of serum VEGF, MMP-2 and MMP-9, ERK and MAPK mRNA levels, and phosphorylated ERK and MAPK protein levels in the uterine tissues than those in model group. CONCLUSIONS: LncRNA BANCR inhibitor can repress the development of ectopic endometrial tissues by inhibiting the generation of angiogenic factors in the EMs focus, and its mechanism may be related to the inhibition on the ERK/MAPK signaling pathway.

MiR-15a-3p suppresses the growth and metastasis of ovarian cancer cell by targeting Twist1.

OBJECTIVE: To investigate the roles of miR-15a-3p in ovarian cancer cell growth and metastasis. PATIENTS AND METHODS: A key role of miR-15a-3p was identified via gene profiling and bioinformatics analysis. The impact of miR-15a-3p on ovarian cancer cell growth, migration and invasion was measured by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT), wound-healing and transwell invasion assays. Bioinformatics and luciferase reporter assays were applied to identify that twist family BHLH transcription factor 1 (Twist1) was the target gene of miR-15a-3p. The miR-15a-3p level and the expression of Twist1 were detected using quantitative Real-time polymerase chain reaction (qRT-PCR) assay. The expressions of N-cadherin and E-cadherin were measured by immunofluorescence staining. Small interfering RNA targeting Twist1 and pCDNA3.1 containing Twist1 were applied to decrease and increase the expression of Twist1, respectively. RESULTS: miR-15a-3p was markedly down-regulated in ovarian cancer. Exogenous up-regulation of miR-15a-3p inhibited the growth, colony formation, migration and invasion of ovarian cancer cell in vitro. Furthermore, a xenograft model indicated that miR-15a-3p inhibited tumour growth and the metastatic potential of ovarian cancer cell in vivo. We found that Twist1 was the direct target of miR-15a-3p in ovarian cancer and that its expression was negatively correlated with the level of miR-15a-3p in ovarian cancer tissues. Up-regulation of miR-15a-3p rescued the inhibitory impact of miR-15a-3p on ovarian cancer cell growth, migration and invasion. Finally, down-regulation of Twist1 mimicked the suppressive effects of miR-15a-3p on ovarian cancer cell. CONCLUSIONS: We demonstrated that miR-15a-3p is down-regulated in ovarian cancer. Up-regulation of miR-15a-3p restrains the growth and metastasis of ovarian cancer cell by regulating Twist1.

Neuroprotective effects of α-lipoic acid on long-term experimental autoimmune encephalomyelitis.

OBJECTIVE: Experimental autoimmune encephalomyelitis (EAE) is an animal model commonly used in research on the acute phase of multiple sclerosis (MS), but studies on the pathology and pathogenesis of EAE with a long disease course are seldom conducted. Besides its antioxidant properties, the comprehensive mechanisms through which α-lipoic acid (LA) affects EAE remain obscure. We here conducted the study to explore the possible mechanisms. MATERIALS AND METHODS: In this study, the following methods were used for investigating the effects of LA on long-term EAE: hematoxylin-eosin staining (HE) and electron microscopic examinations of pathological changes; Western blotting of ß-amyloid precursor protein (ß-APP) and myelin basic protein (MBP); Enzyme-linked immunosorbent assay (ELISA) of tumor necrosis factor-α (TNF-α), transforming growth factor-ß (TGF-ß), superoxide dismutase (SOD), malondialdehyde (MDA) as well as flow cytometry of CD4+CD25+FoxP3+ regulatory T cells (Tregs). RESULTS: The results showed: (1) diverse pathological features of long-term relapsing-remitting EAE; (2) relatively increased MBP and reduced ß-APP expression in LA recipients 180 days after onset; (3) down-regulated TNF-α and up-regulated TGF-ß levels in LA recipients 7 days after onset; (4) lower MDA and higher SOD levels in LA recipients 180 days after onset; (5) increased Treg levels in LA recipients 7 days after onset. CONCLUSIONS: Aside from oxidative stress, LA possessed anti-inflammatory and immunomodulatory effects on EAE. LA might be a promising candidate for MS treatment.

miR-98-TXLNG1 (FIAT)/Sp7 function loop mediates osteoblast mineralization.

OBJECTIVE: To investigate the role of microRNAs (miRNAs) and its mechanism in osteoblast mineralization. MATERIALS AND METHODS: Real-time polymerase chain reaction (PCR), Northern Blot, and Western Blot were used to identify the expression mode of regulators. Overexpression and down-regulation experiments were carried out to study the role of miR-98 and interactions between regulators. Bioinformatics calculation and luciferase reporter assay were used to prove the target gene. Electrophoretic mobility shift assay (EMSA), chromatin immunoprecipitation (CHIP), and promoter luciferase reporter assay confirmed the relationship between the regulator and the promoter of miR-98. RESULTS: MiR-98 was up-regulated during osteoblast mineralization. Overexpression of miR-98 promoted osteoblast mineralization. Factor inhibiting activating transcription factor 4 (ATF4)-mediated transcription (FIAT), a negative regulator of osteoblast differentiation, was confirmed to be a target of miR-98. As a motivator in osteoblast mineralization, Sp7 transcription factor 7 (Sp7) promoted miR-98 transcription by a combination on the promoter region. CONCLUSIONS: Our study showed that miR-98 was an important regulator in osteoblast mineralization and miR-98 carried out its function through a novel miR-98-FIAT/Sp7 regulatory loop. It provides new insights into the roles of miRNAs in osteoblast mineralization.

Long non-coding RNA MALAT1 regulates ovarian cancer cell proliferation, migration and apoptosis through Wnt/ß-catenin signaling pathway.

OBJECTIVE: Long non-coding RNA (LncRNA) MALAT1 is an important regulatory molecule in many diseases, especially in ovarian cancer. We aimed at exploring the function of MALAT1 in ovarian cancer and at clarifying its mechanisms. PATIENTS AND METHODS: The expression level of MALAT1 in ovarian cancer tissues, para-carcinoma tissues and ovarian cancer cell lines were analyzed by Real-time polymerase chain reaction (RT-PCR). The cell proliferation rate was detected by CCK8 assay in SKOV3 and HO8910 cells. Transwell was used to detect the invasion and migration activities in SKOV3 and HO8910 cells. The cell cycle distribution and apoptosis rate were measured by flow cytometry analysis. The expression level of Dvl2, GSK-3ß, ß-catenin and cyclin D1 were detected by RT-PCR and Western blot. RESULTS: The relative expression level of MALAT1 was identified to be aberrantly up-regulated in ovarian cancer tissues and cell lines. The high expression level of MALAT1 was associated with poor prognosis in ovarian cancer patients. The down-regulation of MALAT1 inhibited cell proliferation, invasion and migration, arrested cell cycle progression in S phase and induced cell apoptosis in ovarian cancer cell lines. Meanwhile, the down-regulation of MALAT1 decreased the expression level of DVL2, ß-catenin and cyclin D1 and increased the expression level of GSK-3ß in SKOV3 and HO8910 cells. Moreover, the inhibitory effect of MALAT1 down-regulation in cell invasion and migration was reversed by SKL2001 activating Wnt/ß-catenin signal pathway and enhanced by XAV939 inhibiting Wnt/ß-catenin signal pathway. CONCLUSIONS: MALAT1 was overexpressed in ovarian cancer and associated to the poor prognosis. The down-regulation of MALAT1 inhibited cell proliferation, invasion and migration, arrested cell cycle progression in S phase and induced cell apoptosis by restraining the activation of Wnt/ß-catenin signaling pathway in ovarian cancer cells.

The diagnostic value of three detection techniques of ultrasound in the active stage of Crohn's disease.

OBJECTIVE: The widespread availability of Ultrasound machines and their relatively low cost and functionality make them an attractive tool to use during the treatment of ongoing Crohn's Disease (CD). This study aims at exploring the value of conventional, power Doppler and contrast-enhanced ultrasound during the active stages of CD. PATIENTS AND METHODS: 24 patients in the active stages of Crohn's disease were enrolled in the study. The full, medial and lateral intestinal wall thicknesses and the thickness ratio of medial to lateral intestinal wall of the segmental lesions were measured by conventional ultrasound. The diseased intestinal wall was also examined by power Doppler ultrasound to assign Limberg classification types: 3 cases were Limberg II, 9 cases were Limberg III and 12 cases were Limberg IV type. Importantly, the full and medial thicknesses of the intestinal walls with different Limberg types were compared, and statistically significant differences were found (p<0.05). Finally, images of the diseased segments were taken by contrast-enhanced ultrasound, and the contrast agent bolus arrival, the inflow and the peak enhancement times were calculated in order to be able to distinguish intestinal wall thickness differences according to different Limberg types. RESULTS: Cases with Limberg types III and IV mostly showed total intestinal enhancement, while Limberg II type cases showed mostly medial intestinal enhancement. When comparing the inflow and peak times of contrast-enhanced ultrasound of patients with different Limberg types, the differences found were statistically significant (p<0.05). CONCLUSIONS: This study confirms incrassation of the intestinal wall being the main ultrasonic appearance of active CD. Both power and contrast-enhanced ultrasound are effective tools in the management of active CD: while power ultrasound can be used to carry out Limberg typing, contrast-enhanced ultrasound can analyze and diagnose incrassation segments of the intestinal wall with different disease stages.

Clinical relevance of sHLA-G-mediated with better graft acceptance in early posttransplantation.

The aim of this study was to measure the level of soluble human leukocyte antigen (sHLA-G) in renal transplant patients, to determine the relationship between these levels and the occurrence of acute rejection episodes, and to identify their influence on graft acceptance early posttransplantation. sHLA-G, as measured by an enzyme-linked immunosorbent assay, was significantly increased (P < .01) early posttransplantation (3 months); the other group maintained low levels throughout the study. The latter group displayed a high incidence of acute rejection episodes and a lower clearance of serum creatinine with a longer period for hemoglobin to recover to normal (P < .01). These results suggested that HLA-G participates in the induction of immunologic tolerance in these recipients.

Upregulation of miR-27a contributes to the malignant transformation of human bronchial epithelial cells induced by SV40 small T antigen.

The introduction of the Simian virus 40 (SV40) early region, the telomerase catalytic subunit (hTERT) and an oncogenic allele of H-Ras directly transforms primary human cells. SV40 small T antigen (ST), which forms a complex with protein phosphatase 2A (PP2A) and inhibits PP2A activity, is believed to have a critical role in the malignant transformation of human cells. Recent evidence has shown that aberrant microRNA (miRNA) expression patterns are correlated with cancer development. Here, we identified miR-27a as a differentially expressed miRNA in SV40 ST-expressing cells. miR-27a is upregulated in SV40 ST-transformed human bronchial epithelial cells (HBERST). Suppression of miR-27a expression in HBERST cells or lung cancer cell lines (NCI-H226 and SK-MES-1) that exhibited high levels of miR-27a expression lead to cell growth arrested in the G(0)-G(1) phase. In addition, suppression of miR-27a in HBERST cells attenuated the capacity of such cells to grow in an anchorage-independent manner. We also found that suppression of the PP2A B56γ expression resulted in upregulation of miR-27a similar to that achieved by the introduction of ST, indicating that dysregulation of miR-27a expression in ST-expressing cells was mediated by the ST-PP2A interaction. Moreover, we discovered that Fbxw7 gene encoding F-box/WD repeat-containing protein 7 was a potential miR-27a target validated by dual-luciferase reporter system analysis. The inverse correlation between miR-27a expression levels and Fbxw7 protein expression was further confirmed in both cell models and human tumor samples. Fbxw7 regulates cell-cycle progression through the ubiquitin-dependent proteolysis of a set of substrates, including c-Myc, c-Jun, cyclin E1 and Notch 1. Thus, promotion of cell growth arising from the suppression of Fbxw7 by miR-27a overexpression might be responsible for the viral oncoprotein ST-induced malignant transformation. These observations demonstrate that miR-27a functions as an oncogene in human tumorigenesis.

α4 is highly expressed in carcinogen-transformed human cells and primary human cancers.

A regulator of the protein phosphatase 2A (PP2A), α4, has been implicated in a variety of functions that regulate many cellular processes. To explore the role of α4 in human cell transformation and tumorigenesis, we show that α4 is highly expressed in human cells transformed by chemical carcinogens including benzo(a)pyrene, aflatoxin B(1), N-methyl-N'-nitro-N-nitrosoguanidine, nickel sulfate and in several hepatic and lung cancer cell lines. In addition, overexpression of α4 was detected in 87.5% (74/80) of primary hepatocellular carcinomas, 84.0% (21/25) of primary lung cancers and 81.8% (9/11) of primary breast cancers, indicating that α4 is ubiquitously highly expressed in human cancer. Functional studies revealed that elevated α4 expression results in an increase in cell proliferation, promotion of cell survival and decreased PP2A-attributable activity. Importantly, ectopic expression of α4 permits non-transformed human embryonic kidney cells (HEKTER) and L02R cells to form tumors in immunodeficient mice. Furthermore, we show that the highly expressed α4 in transformed cells or human tumors is not regulated by DNA hypomethylation. A microRNA, miR-34b, that suppresses the expression of α4 through specific binding to the 3'-untranslated region of α4 is downregulated in transformed or human lung tumors. Taken together, these observations identify that α4 possesses an oncogenic function. Reduction of PP2A activity due to an enhanced α4-PP2A interaction contributes directly to chemical carcinogen-induced tumorigenesis.

HLA-B*15 subtypes distribution in Han population in Beijing, China, as compared with those of other populations.

To identify HLA-B*15 subtypes distribution in Han population in Beijing, People's Republic of China, 826 unrelated healthy individuals were typed using the polymerase chain reaction-sequence-based typing method. Within the 246 HLA-B*15 positive individuals, 29 HLA-B*15 alleles were identified, the most predominant of which is B*1501 (40.07%), followed by B*1502 (12.87%), B*1511 (12.87%), B*1518 (9.19%) and B*1532 (3.31%). The distribution of HLA-B*15 subtype frequencies was compared between the Beijing Han, eight other Chinese ethnic minorities and six Chinese populations covering the mainland of China, Taiwan, Hong Kong and Singapore. A neighbor-joining phylogenetic tree was constructed and revealed that the Beijing Han population clustered into the northern populations group and had a closer relationship with northern Han and Hui than with southern Han or other ethnic minorities. These results thus provide useful information that can be used in anthropology, selection for bone marrow transplantation as well as in disease-association study, such as in carbamazepine (CBZ)-induced Stevens-Johnson syndrome and toxic epidermal necrolysis.

Unsuspected Prostate Carcinoma and Prostatic Intraepithelial Neoplasm in Taiwanese Patients Undergoing Cystoprostatectomy.

Although the incidence and death rates for cancer of the prostate (CaP) in Taiwan have been among the lowest in the world, they have increased remarkably in recent years. Because of the very low autopsy rate in this country, prostate specimens obtained via cystoprostatectomy may provide a unique opportunity to study the incidence and status of latent cancer. From January 1992 to December 1997, 49 prostate specimens were obtained from patients with transitional-cell carcinoma of the urinary bladder (48 cases) or pelvic melanoma (one case). Patients' ages ranged from 47 to 89, with a mean age of 67.8 years. No patient had any clinical indication of CaP, as assessed by digital rectal examination. Each prostate was prepared with whole-mount transverse serial sections at 3-mm intervals from the apex to the bladder neck. The stained slides reviewed by two pathologists to evaluate the frequency and pathological status of acinar cancer lesions and high-grade prostatic intraepithelial neoplasia (PIN). Of the 49 patients evaluated, 16 (33%) had evidence of adenocarcinoma, and 24 (49%) had high-grade PIN. The incidence of unsuspected CaP in patients aged 40 to 59, 60 to 69, and >/=70 years was 25%, 32%, and 37%, respectively. The frequency of high-grade PIN in patients aged 40 to 59, 60 to 69, and >/=70 years was 25%, 42%, and 64%, respectively. The incidence of high-grade PIN in the 16 patients with unsuspected CaP was significantly higher than in the 31 patients without this early cancer (75% v 36%). Of the 16 patients with unsuspected cancer, 5 had multiple tumors (3 patients with two and 2 with multiple foci). The mean volume of the 24 tumors was 0.0786 cm(3), with a range of 0.008 to 0.393 cm(3), but only 6 tumors exceeded 0.1 cm(3) in volume (0.112, 0.112, 0.164, 0.245, 0.262, and 0.393 cm(3)). Eighty-eight percent of these early cancers were low grade (Gleason score 2-4). All unsuspected CaP were organ confined. The frequency of unsuspected CaP in Taiwanese men is relatively higher than in Chinese, as previous reported by Dr. Gu. However, the incidence of this latent cancer is comparable to that of U.S. men of the same age. These findings, together with the high incidence of high-grade PIN, suggest that the initial step in the induction of CaP in indigenous Taiwanese is similar to that in U.S. men. The lower number of reports of CaP in Taiwan might be attributable to: (1) lower volume of latent cancer in the Taiwanese compared with U.S. men; (2) underestimation of the incidence rate of CaP in Taiwan; or (3) different genetic or environmental status leading to a different progression rate.