Aberrantly high DEPDC1B expression leads to poor prognosis in patients with lower-grade gliomas.

OBJECTIVE: DEPDC1B, which encodes DEP domain-containing protein 1B, exerts pathogenic effects in diverse cancers, but no such effect has been reported in the case of lower-grade glioma (LGG). Therefore, we sought to investigate the relationship between DEPDC1B expression and the prognosis of patients with LGG and reveal the underlying molecular mechanism. MATERIALS AND METHODS: First, RT-qPCR and immunohistochemical staining were used to examine DEPDC1B mRNA and protein expression in LGG. Second, transcriptomic data were collected from The Cancer Genome Atlas and Chinese Glioma Genome Atlas databases to investigate the impact of DEPDC1B expression on LGG patients by using the Kaplan-Meier survival analysis, receiver operating characteristic analysis and Cox models. Third, the effects of DEPDC1B on LGG cell proliferation and migration were revealed using wound-healing and Cell Counting Kit-8 assays and Ki67 immunofluorescence staining. Fourth, the Tumor Immune Estimation Resource database was used to examine how DEPDC1B affects the LGG immune microenvironment, and gene set enrichment analysis was used to uncover the signaling pathways in which DEPDC1B is involved in LGG. RESULTS: DEPDC1B was significantly upregulated in both LGG cells and tissues, and high expression of DEPDC1B contributed to poor prognosis of LGG patients and represented an independent risk factor for LGG. Moreover, DEPDC1B knockdown reduced the proliferation and migration abilities of LGG cells. Lastly, DEPDC1B was found to be positively associated with multiple immune infiltrates and immune-checkpoint markers. CONCLUSIONS: Our findings indicate for the first time that DEPDC1B is a pathogenic gene in LGG. More importantly, we provide a new biomarker and immunotherapeutic target for improving the diagnosis and treatment of LGG patients.

ILF3-AS1 promotes the aerobic glycolysis and proliferation of melanoma cells by regulating miR-493-5p/PDK1 pathway.

BACKGROUND: The aim of this study was to investigate the role of ILF3-AS1 in regulating the survival of melanoma and its molecular mechanism. METHODS: The relative expression level of ILF3-AS1 in melanoma was assessed by qPCR. The effect of ILF3-AS1 and PDK1 on the cell viability was tested by MTT assay. Glucose uptake colorimetric assay, lactate assay, the measurements of extracellular acidification rate (ECAR) and Oxygen consumption rate (OCR) were performed to test the effect of ILF3-AS1 and PDK1 on the cellular glycolysis. Luciferase assay was conducted to detect the interactions of ILF3-AS1, miR-493-5p and PDK1. RNA immunoprecipitation chip (RIP) assay was used to detect the enrichments of ILF3-AS1 and miR-493-5p in the complex. Protein level of PDK1 was detected by western blot analysis. RESULTS: qPCR revealed that ILF3-AS1 was upregulated in human melanoma cell lines. MTT assay showed that ILF3-AS1 knockdown blunted cell proliferation, which was rescued by the overexpression of PDK1. Glucose uptake colorimetric assay, lactate assay, the measurements of ECAR and OCR indicated that ILF3-AS1 promoted glycolysis through PDK1. Western blotting results showed that ILF3-AS1 overexpression promoted PDK1 expression, which was prevented by miR-493-5p overexpression in SK-MEL-1 cells. CONCLUSIONS: ILF3-AS1 promotes the aerobic glycolysis and survival of melanoma cells involving miR-493-5p/PDK1 pathway.

Photoluminescence and magnetism integrated multifunctional black phosphorus probes through controllable PO bond orbital hybridization.

Biological probes with integrated photoluminescence and magnetism characteristics play a critical role in modern clinical diagnosis and surgical protocols combining fluorescence optical imaging (FOI) with magnetic resonance imaging (MRI) technology. However, traditional magnetic semiconductors can easily generate a spin splitting at the Fermi level and half-metallic electronic occupation, which will sharply reduce the radiation recombination efficiency of photogenerated carriers. To overcome this intrinsic contradiction, we propose a controllable oxidation strategy to introduce some particular PO bonds into black phosphorus nanosheets, in which the p orbital hybridization between P and O atoms not only provides some carrier recombination centers but also leads to a room-temperature spin polarization. As a result, the coexistence of photoluminescence and magnetism is realized in multifunctional black phosphorus probes with excellent biocompatibility. This work provides a new insight into integrating photoluminescence and magnetism together by intriguing atomic orbital hybridization.

MicroRNA-1286 inhibits osteogenic differentiation of mesenchymal stem cells to promote the progression of osteoporosis via regulating FZD4 expression.

OBJECTIVE: The aim of this study was to investigate whether microRNA-1286 could inhibit the osteogenic differentiation of human marrow mesenchymal stem cells (hMSCs) by regulating FZD4 expression and promoting the progression of osteoporosis. PATIENTS AND METHODS: Quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) was used to detect the expression of microRNA-1286 in the serum of patients with osteoporosis. Meanwhile, microRNA-1286 expression in different stages of osteogenic differentiation of hMSCs was measured by qRT-PCR as well. After overexpression of microRNA-1286 and FZD4 in hMSCs, the mRNA expression levels of microRNA-1286, alkaline phosphatase (ALP), RUNX2 and osteocalcin (OCN) were detected by qRT-PCR. The protein expression levels of RUNX2 and OCN were detected by Western blot. Meanwhile, alkaline phosphatase (ALP) activity and expression in cells were examined using ALP assay kit and ALP staining method, respectively. Cell mineralized nodules were detected through the alizarin red staining test. Bioinformatics method was used to predict the binding site of microRNA-1286 to FZD4. Subsequent luciferase reporter gene assay was performed to verify whether microRNA-1286 could combine with FZD4. After overexpression or knockdown of microRNA-1286, the mRNA and protein expressions of FZD4 were analyzed using qRT-PCR and Western blot assay, respectively. After the simultaneous overexpression of microRNA-1286 and FZD4 in hMSCs, the mRNA expression levels of ALP, RUNX2 and OCN, ALP activity and content, and cell mineralization ability were successively examined. RESULTS: The expression of microRNA-1286 in the serum of patients with osteoporosis was significantly higher than that of the normal population. Meanwhile, microRNA-1286 expression decreased with the increase of osteogenic differentiation days of hAMSCs. After the overexpression of microRNA-1286, ALP, RUNX2, and OCN levels, ALP activity, RUNX2, and OCN protein levels, as well as mineralized nodule formation were significantly reduced. However, results were reversed when FZD4 was simultaneously up-regulated. Luciferase reporter gene assay results verified that microRNA-1286 could bind to FZD4. After the overexpression of microRNA-1286, the mRNA and protein expressions of FZD4 were found significantly down-regulated. However, results were reversed after knocking down microRNA-1286. Furthermore, the simultaneous overexpression of microRNA-1286 and FZD4 could counteract the inhibitory effect of over-expression of microRNA-1286 on osteogenic differentiation of hMSCs. CONCLUSIONS: MicroRNA-1286 can regulate FZD4 expression and inhibit osteogenic differentiation of hMSCs, thereby promoting the development of osteoporosis.

Clock mutant promotes osteoarthritis by inhibiting the acetylation of NFκB.

OBJECTIVES: To examine the effect of the circadian gene Clock on posttranscriptional function and pro-inflammatory mechanisms in osteoarthritis (OA). METHODS: The cartilage from Clock mutant mice was assessed using histology, (OA) score, and real-time polymerase chain reaction (PCR) quantification of key pro-inflammatory genes. Nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB) translocation, posttranslational state and expression levels during day and night conditions were assessed using immunoblot and IP. The regulation of transcription by Clock in cartilage tissue was assessed by using chromatin immunoprecipitation (ChIP) and luciferase assays. Total acetylation level and pattern over 24 h were quantified using immunoblot and real-time PCR. Finally, the effects of exogenous Clock nanoparticle treatment were quantified by histology and immunoblot. RESULTS: The Clock mutation significantly promoted the degradation of cartilage and the expression of the key pro-inflammatory mediators, IL-1ß, IL-6 and MCP-1. The Clock mutation significantly promoted NFκB nuclear translocation. The circadian protein CLOCK positively regulates NFκB at the transcriptional level by binding the E-box domain. The Clock mutation significantly inhibited the total lysine acetylation level in cartilage and inhibited NFκB acetylation at the Lys310 residue but promoted phosphorylation at the Ser276 residue. The forced expression of Clock in vivo inhibited NFκB activation by increasing acetylation and decreasing phosphorylation levels and by decreasing cartilage damage and inflammation. CONCLUSIONS: This study demonstrates the mutation of Clock promotes inflammatory activity by mediating the posttranscriptional regulation of NFκB in OA pathogenesis.

[Resting state fMRI study of emotional network in patients with postconcussion syndrome].

Objective: To discuss functional connectivity changes in the emotional network of patients with post-concussion syndrome (PCS) and their clinical significance by resting-state functional magnetic resonance imaging (rs-fMRI). Methods: Twenty-seven patients with PCS were recruited from the Department of Neurosurgery of Anhui provincial hospital affiliated to Anhui medical university from October 2015 to April 2016, and 27 healthy subjects were recruited as the controls. The Hamilton Anxiety Scale (HAMA) and The Hamilton Depression Scale (HAMD) were used to evaluate the emotional state of two groups of subjects. All fMRI data were preprocessed after RS-fMRI scanning, the left and right amygdala were selected as region of interest (ROI) to make functional connectivity (FC) calculation with the whole brain and then the results were did statistical analysis in order to obtain the altered brain areas of amygdala and whole brain functional connectivity in the PCS patient, to understand the functional changes of emotional network. Results: HAMA and HAMD scores of PCS group and the health controls had significant statistical difference (HAMA: the PCS group 9.8±1.5, the health controls 4.5±1.2, P=0.044; HAMD: the PCS group 12±1.2, the health controls was 4.2±1.5, P=0.024). Compared with the health controls, the left amygdala in PCS patients showed decreased FC with left insula, left putamen, left anterior cingulate gyrus, left inferior orbital frontal gyrus, left medial superior frontal gyrus, bilateral superior temporal gyrus, left superior temporal pole, bilateral supramarginal gyrus et al, on the contrary with the increased FC with right superior orbital frontal gyrus, right middle frontal lobe, right orbital frontal lobe, right middle frontal gyrus. The right amygdala in PCS patients showed decreased FC with bilateral putamen, right inferior orbital frontal gyrus, left insula, bilateral precuneus, bilateral superior temporal pole, right superior temporal gyrus, right supramarginal gyrus et al. Similarly, it had the increased FC with the left middle occipital lobe and the left superior occipital lobe. Conclusion: Abnormal emotional network function of PCS patients in resting state, which may be one of the reasons that lead to emotional and cognitive dysfunction in PCS patients.

TGF-ß1 promotes human hepatic carcinoma HepG2 cells invasion by upregulating autophagy.

OBJECTIVE: To study the role of TGF-ß1 in autophagy and invasion ability of human hepatic carcinoma HepG2 cells. MATERIALS AND METHODS: Cultured HepG2 cells were treated with different concentrations of TGF-ß1 for 24 h. The protein expression levels of autophagy relative marker LC3 and Beclin1 were detected by Western blot. The effect of TGF-ß1 on invasion ability of HepG2 cells was detected with transwell method. RESULTS: The results demonstrated that TGF-ß1 was able to activate autophagy of HepG2 cells in a dose-dependent manner. Autophagy inhibitor 3-methyladenine (3-MA) could reverse TGF-ß1 induced autophagy process. Also, TGF-ß1 significantly promotes the invasion ability of HepG2 cells; however, this process could effectively reverse by autophagy inhibitor 3-MA. CONCLUSIONS: TGF-ß1 enhances HepG2 cells invasion by upregulating autophagy.

Identification of IgE and IgG1 specific antigens in Echinococcus granulosus cyst fluid.

Cystic echinococcosis (CE) is an anthropozoonotic disease with worldwide distribution and is caused by the cestode Echinococcus granulosus. Anaphylactic shock induced by CE rupture is a serious complication especially in patients with hydatid infections, as the resulting leakage of fluid contains highly toxic endogenous antigen. We aimed to isolate and identify the antigens of specific IgE and IgG1 (sIgE and sIgG1) in E. granulosus cyst fluid (EgCF). Crude antigen for EgCF was prepared from E. granulosus-infected sheep liver. Antigens were separated and identified by one-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (1D SDS-PAGE), two-dimensional gel electrophoresis (2-DE), and immunoblotting. Results of 1D SDS-PAGE and immunoblotting showed that 40.5 kDa protein was the major antigen of sIgE, and 35.5 kDa protein was the major antigen of sIgG1 in EgCF. Results of 2-DE and immunoblotting showed that main antigens of sIgE in EgCF were four proteins with pI values ranging from 6.5 to 9.0 and a molecular weight of 40.5 kDa. Main antigens of sIgG1 in EgCF were five proteins with pI values ranging from 6.5 to 9.0 and a molecular weight of 35.5 kDa. The antigens identified for sIgE and sIgG1 can provide critical insights into cellular and molecular mechanisms underlying anaphylactic shock induced by CE.

[Evaluation of endometriosis fertility index in follow-up treatment of endometriosis combined with infertility patients after laparoscopic surgery].

Objective: To explore the application of endometriosis fertility index (EFI) in guidance after laparoscopic surgery of endometriosis patients combined with infertility and to explore methods to improve pregnancy rate in different EFI groups. Methods: A prospective research was done in endometriosis patients combined with infertility in Beijing Obstetrics and Gynecology Hospital from January 2010 to June 2011, after laparoscopic surgery, these 146 patients were divided into 3 groups by EFI score. Using different pregnancy guidance, these patients had 5 years follow-up. Results: (1) The 5 years overall pregnancy rate was 89.0% (130/146). The pregnancy rate was 95.7% (45/47) in EFI≥9 group, 92.8% (77/83) in EFI 5-8 group and 8/16 in EFI≤4 group, three groups were all reach satisfactory pregnancy rate; the rate of the first two groups had no statistically significance (P=0.498), but had significant difference with the last group (P<0.01). (2) In EFI≥5 patients, pregnancy rate was the highest in 6 months after operation; in EFI≥9 group, the pregnancy rate was 66.7% (30/45), and EFI 5-8 group was 50.6% (39/77). (3) EFI≥9 group had the highest natural pregnancy rate [83.6% (46/55)], natural pregnancy rate was significant statistical different in different EFI groups (P=0.001). Conclusions: EFI score is a useful evaluation in predicting and guiding pregnancy in endometriosis patients combined with infertility after laparoscopic surgery. EFI score guidance, strict post-operation management and positive pregnancy scheme could significantly improve the pregnancy rate of endometriosis patients with infertility.

Identification of IgE and IgG1 specific antigens in Echinococcus granulosus cyst fluid

Cystic echinococcosis (CE) is an anthropozoonotic disease with worldwide distribution and is caused by the cestode Echinococcus granulosus. Anaphylactic shock induced by CE rupture is a serious complication especially in patients with hydatid infections, as the resulting leakage of fluid contains highly toxic endogenous antigen. We aimed to isolate and identify the antigens of specific IgE and IgG1 (sIgE and sIgG1) in E. granulosus cyst fluid (EgCF). Crude antigen for EgCF was prepared from E. granulosus-infected sheep liver. Antigens were separated and identified by one-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (1D SDS-PAGE), two-dimensional gel electrophoresis (2-DE), and immunoblotting. Results of 1D SDS-PAGE and immunoblotting showed that 40.5 kDa protein was the major antigen of sIgE, and 35.5 kDa protein was the major antigen of sIgG1 in EgCF. Results of 2-DE and immunoblotting showed that main antigens of sIgE in EgCF were four proteins with pI values ranging from 6.5 to 9.0 and a molecular weight of 40.5 kDa. Main antigens of sIgG1 in EgCF were five proteins with pI values ranging from 6.5 to 9.0 and a molecular weight of 35.5 kDa. The antigens identified for sIgE and sIgG1 can provide critical insights into cellular and molecular mechanisms underlying anaphylactic shock induced by CE.

[Prevention strategies for surgical complications of sphenoid wing meningiomas].

OBJECTIVE: To explore the prevention strategies for surgical complications of sphenoid wing meningiomas. METHODS: A total of 76 cases of sphenoid wing meningiomas were included from The People's Hospital of Henan Province from 2011 to 2014. Based on the preoperative imaging data, the patients were divided into 28 cases of lateral type, 16 cases of middle type and 32 cases of medial type. According to the different characteristics of different types of sphenoid wing meningiomas, appropriate preoperative, intraoperative and postoperative measures to prevent complications were carried out. RESULTS: A total of 2 cases of facial nerve injury occurred in 28 cases with lateral type; 1 case of intraoperative subarachnoid hemorrhage and brain swelling occurred in 16 cases with middle type; 3 cases of oculomotor nerve damage, 1 case of optic nerve injury and 4 cases of vascular injury occurred in 32 cases with the medial type. The total incidence of complications was 14.5% (11/76). CONCLUSION: Detailed preoperative planning according to the characteristics of different types of sphenoid wing meningiomas, intraoperative use of microsurgery and Cavitron Ultrasonic Surgical Aspirator (CUSA), following the principle of in situ removal, controlling the extent of resection of tumor adhering to cavernous sinus and the great vessels, the severe surgery complications of sphenoid wing meningiomas can be reduced.

Adjudin--A Male Contraceptive with Other Biological Activities.

BACKGROUND: Adjudin has been explored as a male contraceptive for the last 15 years since its initial synthesis in the late 1990s. More than 50 papers have been published and listed in PubMed in which its mechanism that induces exfoliation of germ cells from the seminiferous epithelium, such as its effects on actin microfilaments at the apical ES (ectoplasmic specialization, a testis-specific actin-rich anchoring junction) has been delineated. OBJECTIVE: Recent studies have demonstrated that, besides its activity to induce germ cell exfoliation from the seminiferous epithelium to cause reversible infertility in male rodents, adjudin possesses other biological activities, which include anti-cancer, anti-inflammation in the brain, and anti-ototoxicity induced by gentamicin in rodents. Results of these findings likely spark the interest of investigators to explore other medical use of this and other indazole-based compounds, possibly mediated by the signaling pathway(s) in the mitochondria of mammalian cells following treatment with adjudin. In this review, we carefully evaluate these recent findings. METHODS: Papers published and listed at www.pubmed.org and patents pertinent to adjudin and its related compounds were searched. Findings were reviewed and critically evaluated, and summarized herein. RESULTS: Adjudin is a novel compound that possesses anti-spermatogenetic activity. Furthermore, it possesses anti-cancer, anti-inflammation, anti-neurodegeneration, and anti-ototoxicity activities based on studies using different in vitro and in vivo models. CONCLUSION: Studies on adjudin should be expanded to better understand its biological activities so that it can become a useful drug for treatment of other ailments besides serving as a male contraceptive.

Model simulation of inflow water to the Baltic Sea based on ¹²9I.

The semi-enclosed Baltic Sea represents a vital economic and recreational resource for more than 90 million people inhabiting its coasts. Extensive contamination of this sea by a variety of anthropogenic pollutants has raised the concern of the people in the region. Quantifying seawater inflow is crucial for estimating potential environmental risks as well as to find the best remedial strategy. We present here a model to estimate water inflow from the North Sea to the Baltic Sea by utilizing ¹²9I as a tracer. The results predicted inflow range of 230-450 km³/y with best fit value around 330 km³/y from the North Sea to the Baltic Sea during 1980-1999. Despite limited time series data on ¹²9I, the model presented here demonstrates a new management tool for the Baltic Sea to calculate inflow water compared to conventional methods (such as salinity, temperature and hydrographic models).

Wenshen Xiaozheng Tang suppresses the growth of endometriosis with an antiangiogenic effect.

OBJECTIVE: To evaluate the effect of Wenshen Xiaozhng Tang (WXT) on ectopic endometrial growth and on angiogenesis in endometrial implants in a rat model. METHODS: Sprague-Dawley rats with endometriotic implants were randomly treated with low-dose WXT, high-dose WXT, or vehicle (negative control) for 28 days. Cell proliferation and vascular density in the lesions were assessed by immunohistochemistry. The levels of VEGF in peritoneal fluid were determined by ELISA. The mRNA expression of HIF-1α and Flk-1 in the endometriotic lesions was evaluated by real-time PCR. RESULTS: WXT treatment significantly decreased the lesion size and inhibited cell proliferation in the endometriotic lesions. Lowered vascular density and reduced mRNA expression of HIF-1α in the endometriotic lesions, associated with decreased concentration of VEGF in peritoneal fluid, were also observed in WXT-treated rats. CONCLUSIONS: These results suggest that WXT could be effective to suppress the growth of endometriosis, partially through its antiangiogenic activity.

Clinicopathological significance of cyclooxygenase-2 and cell cycle-regulatory proteins expression in patients with esophageal squamous cell carcinoma.

The aim of this study was to examine the expression of the molecular markers cyclooxygenase-2 (COX-2), Ki-67, cyclin A, and p27 in patients with esophageal squamous cell carcinoma (ESCC), to ascertain the relationship of these makers with the clinicopathological significance of the patients, and to assess the additional prognostic value of the expression profile of these proteins for ESCC patients. The expression levels of COX-2, Ki-67, cyclin A, and p27 proteins of a series of primarily resected ESCC samples were determined by immunohistochemistry method. Clinicopathological and molecular factors affecting survival were analyzed by multivariate analysis. A total of 78 specimens were included in this study. Expression of COX-2 was observed in 43 (55.1%) cases, and high levels of expression of Ki-67, p27, and cyclin A were observed in 57 (73.0%), 33 (42.3%), 43 (55.1%) cases, respectively. The results of univariate survival analysis indicated that more advanced tumor stage, lymph node involvement, systemic dissemination, the levels of expression of COX-2, Ki-67, cyclin A, and p27 were associated with survival (all P-value < 0.05). Multifactorial survival analysis revealed that only lymph node involvement, over-expression of cyclin A, and low p27 expression were associated with the survival of the patients (hazard ratios = 2.83, 4.7, 2.9, respectively; P= 0.025, 0.042, 0.005, respectively). Among the molecular markers assessed, the expression of cell proliferation markers cyclin A and p27 are independent prognostic factors in patients with ESCC, whereas neither COX-2 nor Ki-67 is of independent prognostic value.

IL-1 receptor antagonist gene polymorphism in idiopathic recurrent spontaneous abortion in a Chinese Han population.

Interleukin-1 receptor antagonist (IL-1Ra) has been supposed to play important roles in pregnancy. The purpose of this study was to evaluate the association between the polymorphisms of IL-1Ra gene (IL1RN) variable number tandem repeat (VNTR) in intron 2 with idiopathic recurrent spontaneous abortion (RSA). Ninety-two RSA patients and hundred normal women with at least one live birth and no history of miscarriage were included in the study. Frequencies of the IL1RN alleles and genotypes were determined. Data revealed that the prevalence of IL1RN allele and genotype was not significant between the RSA and control group (all P > 0.05). Our finding indicated that the polymorphism VNTR of IL1RN gene in intron 2 may not be a risk factor for RSA in the Chinese Han population.

MicroRNA-320 expression in myocardial microvascular endothelial cells and its relationship with insulin-like growth factor-1 in type 2 diabetic rats.

1. The aim of the present study was to determine the role of myocardial microvascular endothelial cells (MMVEC) in impaired angiogenesis of type 2 diabetic Goto-Kakizaki (GK) rats. 2. A microRNA (miRNA) microarray was used to assess miRNA expression in MMVEC from GK and Wistar rats. Upregulation of miRNA-320 was observed in MMVEC from GK rats using real-time reverse transcription-polymerase chain reaction (RT-PCR). 3. So far, nine miRNAs have been reported to target angiogenic factors and/or receptors, including kinase insert domain containing receptor (Flk-1), insulin-like growth factor 1 (IGF-1) and insulin-like growth factor 1 receptor (IGF-1R). The predicted genes targeted by miR-320 include Flk-1, IGF-1 and IGF-1R. Western blot analysis and RT-PCR were used to analyse the protein and mRNA expression, respectively, of the putative genes IGF-1 and IGF-1R. The expression of IGF-1 and IGF-1R proteins decreased significantly in diabetic MMVEC. However, the expression of IGF-1 mRNA increased rather than decreased. The mRNA expression of IGF-1R did not differ significantly between diabetic and control MMVEC. 4. Transfection of an miR-320 inhibitor into MMVEC from GK rats confirmed that miR-320 impaired angiogenesis. The proliferation and migration of diabetic MMVEC improved after transfection of the miR-320 inhibitor. In addition, the miR-320 inhibitor significantly increased the expression of IGF-1 protein, but had no effect on the expression of IGF-1R. 5. Eleven miRNAs were upregulated in MMVEC from GK rats compared with those in Wistar rats: let-7e, miR-129, miR-291-5p, miR-320, miR-327, mir-333, miR-363-5p, miR-370, miR-494, miR-503 and miR-664. 6. The results indicate that upregulation of miR-320 in MMVEC from GK rats may be responsible for the inconsistency between the expression of IGF-1 protein and mRNA and therefore related to impaired angiogenesis in diabetes. Transfection of an miR-320 inhibitor may be a therapeutic approach for the treatment of impaired angiogenesis in diabetes.

Effects of high glucose plus high insulin on proliferation and apoptosis of mouse endothelial progenitor cells.

OBJECTIVE: To study the effects of high glucose plus high insulin (GI) on proliferation, apoptosis, morphology and differentiation of endothelial progenitor cells (EPC). METHODS: EPC were isolated and identified by magnetic cell sorting and flow cytometry. EPC proliferation and apoptosis were measured by MTT assay and Annexin-V/PI double labeling, respectively. Cell proliferation- and apoptosis-related factors were examined by Western blotting. RESULTS: Seven days after GI treatment, EPC were arrested at the G(0)/G(1) phase. Treatment with high glucose alone (HG) or GI but not HI (high insulin alone) decreased EPC proliferation and their expression of cyclin E, cdk2 and PCNA compared to control. The inhibitory effects of HG on EPC proliferation and growth-related proteins were more significant than those of GI. HG, GI and HI promoted EPC apoptosis along with caspase-3 overexpression. CONCLUSION: The result indicated that GI-induced apoptosis and growth inhibition might be one of mechanisms of EPC reduction and dysfunction in diabetes.

Characterization of OmpK, GAPDH and their fusion OmpK-GAPDH derived from Vibrio harveyi outer membrane proteins: their immunoprotective ability against vibriosis in large yellow croaker (Pseudosciaena crocea).

AIM: To investigate the immunoprotection of three recombinant proteins derived from the Vibrio harveyi outer membrane proteins (OMPs) OmpK, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and their fusion OmpK-GAPDH as vaccine candidates from vibriosis of large yellow croaker (Pseudosciaena crocea). METHODS: The ompK gene, of which the leader sequence was omitted, was fused with the gapdh gene. Three recombinant proteins r-OmpK, r-GAPDH and r-OmpK-GAPDH were expressed and purified. Western blots were carried out to detect the specificity of the antibodies raised against the recombinant proteins; Fish were immunized with recombinant proteins and challenged by native V. harveyi. The immunoresponse to the recombinant proteins were determined by ELISA and phagocytic activity assay. CONCLUSIONS: The fusion protein r-OmpK-GAPDH can afford greater protection against the wild V. harveyi than r-OmpK or r-GAPDH alone or their mixture in humoral and cellular immunity, indicating that OmpK and GAPDH could produce a synergistic immunoprotection against vibriosis of large yellow croaker (Pseudosciaena crocea) when fused into OmpK-GAPDH with a linker. SIGNIFICANCE AND IMPACT OF THE STUDY: It has been realized that a multi-component OMP antigen can induce a higher frequency of immune effectors than a single OMP. The results presented here bring forth a good suggestion for the subunit vaccine design based on the OMPs of gram-negative pathogens.