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Previous reports have indicated that failure in cloning monkey is attributed to the removal of nuclear mitotic apparatus (NuMA) during enucleation and subsequent abnormal organization of mitotic apparatus. This study investigated the transformation and assembly of tubulin and NuMA protein during the first cell cycle of cloned monkey embryos reconstructed by using enucleated rabbit oocytes as recipients. After the oocyte fused with a fibroblast, extensive microtubule organization was observed around the introduced nucleus in most reconstructed embryos, suggesting the introduction of a somatic cell centrosome. A high proportion of fibroblast nuclei transferred into non-activated oocytes underwent premature chromosome condensation (PCC), transient spindle organization and chromosomes separation, followed by the formation of two pronucleus-like structures. In contrast, fibroblast nuclei in pre-activated ooplasm rarely underwent PCC, but formed a swollen pronucleus-like structure. Normal spindles were observed in about one third of the cloned embryos reconstructed by both methods. After transferring monkey fibroblasts into NuMA-removed enucleated rabbit oocytes, NuMA was localized in pseudo-pronuclei and gradually moved to mitotic spindle poles at the first mitotic spindle poles. NuMA antibody microinjection resulted in spindle disorganization and chromosome misalignment, but did not significantly affect early cleavage. Our findings indicate that: 1. NuMA in donor monkey fibroblast may contribute to form a normal spindle in enucleated rabbit oocyte; 2. when non-activated cytoplasts and pre-activated cytoplasts are used as recipients, the donor nuclei undergo different morphological changes, but yield similar early embryo development; 3. although abnormal spindle organization and chromosome alignment may cause low efficiency of animal cloning, these abnormalities do not significantly affect early cleavage.

AIM: To investigate the effect of probiotics supplemented by gut on the tight junctions of epithelial cells, barrier function and the microflora of rats with abdominal infection. METHODS: After the model of cecal ligation and perforation established, SD rats were divided into two groups: parenteral nutrition (PN) group and PN+probiotics (probiotics) group, PN solution was supplemented by neck vein and probiotics was delivered via the jejunostomy tube for five days. Vena cava blood and the homogenated tissue of liver, lung and mesenteric lymph nodes were cultured to determine the bacterial translocation rate (BTR). The ultra-structure of epithelial tight junctions and microvilli of the gut were observed by electron microscopy; occluding expression was measured by indirect-immune fluorescence method; anaerobic bacterial growth by anaerobic culture and DNA fingerprint of bacterial colonies of the feces by PCR. RESULTS: The quantity of lactobacteria and bifydobacteria in probiotics group was higher than that of PN group. The profiles of DNA fingerprint expression in probiotics group were similar to that in the normal group, a new 16S rDNA sequence appeared in the profile in PN group. The occludin expression, the integrality of the gut epithelial tight junction and microvilli in probiotics group were improved as compared with PN group. The BTR and endotoxin in blood were reduced more significantly in probiotics group as compared with PN group. CONCLUSION: The probiotics could improve the gut microflora disturbance, increase occludin expression, maintain the gut epithelial tight junction and decrease the bacterial translocations rate.

Objective: To study the expressions of basic fibroblast growth factor (bFGF) and its receptor (bFGFR) in bone marrow of mice with acute radiation injury, and to evaluate the effect of Ligustrazine (Lt) on them. Methods: Fifty-six Kunming mice of clean grade were randomly divided into 3 groups, the normal group, the control group and the Lt group. Mice in the latter two groups were once homogeneously systemic irradiated with 6.0 Gy of 60 Co, with the absorption dose rate of 0. 56 Gy/min, then treated with saline (0.2 ml/ mice) or Lt (2 mg/mice) respectively, twice a day through gastrogavage for successive 13 days. Mice were sacrificed in batch on the 3rd, 7th and 14th day by cervical dislocation to collect the bilateral femoral bone marrow for preparing bone marrow mono-nuclear cell (BMMNC) suspension. The bFGFR expression on surface of BMMNC was determined by flow cytometry; and the bFGF expres-sion level in one side of femoral bone marrow tissue was detected by immunohistochemistry with SABC-AP assay. Results: The bFGF expression in bone marrow of mice on the 3rd, 7th and 14th day after acute radiation injury all were significantly lower than that of the normal mice (P0.05 or P0.01). The expressions of bFGF and bFGFR in the Lt group detected were significantly higher than that in the control group detected at the corresponding time points (P0.05 or P 0.01). Conclusion:By way of enhancing bFGF expression in bone marrow and bFGFR expression on surface of BMMNC to accelerate the repairing of hemopoietic micro-environment in bone marrow might be one of the mechanisms of Lt in promoting hemopoietic function reconstitution after acute radiation injury.Original article on CJITWM (Chin) 2004;24(5):439

AIM: To detect the genetic alteration and abnormal expression of cyclin D1 in gastric carcinoma and investigate its clinicopathologic significance in advanced gastric carcinoma. METHODS: Proteins of cyclin D1 were detected by immunohistochemistry in 42 cases of advanced gastric carcinoma with their follow-up data available, 27 cases of early stage carcinoma, 21 cases of gastric adenoma, 22 cases of hyperplastic polyp and 20 cases of normal mucosa adjacent to adenocarcinomas. Genetic alteration of cyclin D1 was detected by Southern blot and expression of cyclin D1 mRNA was detected by PT-PCR in 42 cases of advanced gastric carcinoma. RESULTS: Cyclin D1 protein was not expressed in normal mucosa, hyperplastic polyp and gastric adenoma, while it was only positively expressed in gastric carcinoma. The expression rate of cyclin D1 protein in early stage gastric carcinoma, advanced gastric carcinoma and lymph node metastasis was 48.1%, 47.4% and 50.0%, respectively. The amplification of cyclin D1 gene was detected in 16.6% of advanced gastric carcinomas. The overexpression of cyclin D1 mRNA was detected in 40.5% of the samples. There was no significant correlation between cyclin D1 protein expression and age, lymph-node metastasis and histological grading in patients with advanced gastric carcinoma (χ 2 = 0.038,0.059,0.241, P > 0.05). Significant correlation was observed between the expression of cyclin D1 protein and the 5-year survival rate (χ 2 = 3.92, P < 0.05). CONCLUSION: Detection of cyclin D1 protein by immunohistochemistry may be useful in the diagnosis of early gastric carcinomas. Patients with positive expression of cyclin D1 protein tend to have a worse prognosis.

AIM: To determine whether expression of the tumor suppressors p16 and Rb is altered in gastric carcinoma. METHODS: Mucosal biopsies were endoscopically obtained from patients with superficial gastritis (n = 12), atrophic gastritis (n = 15), atypical hyperplasia (n = 20) and gastric cancer (n = 40). Upon obtainment, all samples were immediately fixed with 10% buffered formalin, embedded in paraffin, and sectioned serially. Protein expression of p16 and Rb was detected by immunohistochemistry (ABC method). RESULTS: The gastric epithelium samples showed various degrees of nuclear immunostaining for p16 and Rb according to the different stage of lesion. Progressive pathology of the lesions was associated with a decreasing trend in positive immunostaining for p16 protein (83.3% > 73.3% > 30.0% > 27.5%) but an increasing trend for Rb protein (25.0% > 46.7% > 60.0% > 67.5%). A negative correlation was found between these two parameters and gastric cancer. Correlation analysis of the 40 cases of gastric cancer identified a negative correlation for 20 of the cases. When positive (n = 9) and negative tissues (n = 11) were compared, a statistically significant difference was found (50.0%, 22.5%, 27.5%) (P < 0.05). CONCLUSION: Abnormal expression of p16 and Rb may play an important role in gastric carcinogenesis.

The administration of MPTP to man and monkey has been shown to cause a neurotoxic effect on the nigrostriatal dopamine system. MPTP was injected in C57-BL black mice, 36 mg per kg for 7 days, which resulted in permanent reduction of dopamine and serotonin levels in the striatum. In the mice pretreated with PLG, although the striatal dopamine level was also reduced, mean dopamine and serotonin levels were significantly higher than in mice given MPTP alone. It is concluded that PLG could protect at least partially the neurotoxic effect of MPTP.

An immunoluminometric assay for plasma lactoferrin has been developed and used to study the levels in children and neonates with viral and bacterial infections. The reference range for plasma lactoferrin was 50–250 μg/l. Lactoferrin levels were significantly higher in patients with bacterial versus viral infections.

The present study assessed the urinary protein with SDS-PAGE technique and by using gentamicin as a positive control for a comparative study to evaluate the renal toxic effect of gossypol. Results indicated that gentamicin could induce proteinuria by alteration of glomerular permeability to cause over-filtration of high molecular weight proteins, damage the brush border (BB) membrane of proximal renal tubules and impair tubular reabsorption function. Gossypol could also induce proteinuria in certain cases of guinea pigs but not at all in gossypol-treated rats. However, an increase in low molecular weight protein bands (M.W. range 20–30kd) in urinary electrophoregram of gossypol-treated rats were detectable.

ObjectiveTo study the influence of Jiang Tang Fang Long formula on insulin production and function in an animal model of diabetic hearing loss.MethodsWistar rats (n=60) were randomly divided into 6 groups (10 in each) to receive no treatment (the normal control, Group A), or to receive intra-peritoneal 55 mg/kg streptozotocin with (Groups C, D and E) or without (Group B) subsequent Jiang Tang Fang Long formula treatment at various doses or Yu Long Wan treatment (Group F). After 60 days, fasting blood glucose (FBG), body weight (BW) and fasting insulin (FINS) were recorded and the HOMA-IR and HOMA-β calculated. Insulin expression in pancreatic tissues was measured by radioimmunoassay.ResultsCompared with animals that received streptozotocin without rescue treatment (Group B), animals that received higher doses of Jiang Tang Fang Long formula(Groups D and E) showed improved indices of diabetes manifestation (P< 0.05) and improved HOMA-β (P0.05).ConclusionOur results suggest that Jiang Tang Fang Long formula may improve pancreatic β-cells function which may explain its efficacy in treating diabetic hearing loss.

In China, at least 9 medical centers are carrying out the studies on factor VIII, especially on hemophilia A. A rabbit antiserum to human VIIIR:Ag was prepared in 1980 and 661 cases were studied with the antiserum, comprising 213 cases of hemorrhagic diseases and 448 cases of normal pregnant women and non-hemorrhagic diseases. The results were similar to those reported in medical literatures. VIIIR:Ag increased progressively with the evolution of the pregnancy reaching up to 230 percent at the third trimester, very close to that observed in toxemia of pregnancy, while the blood from umbilical cord had a normal level of VIIIR:Ag. Studies on the localization of VIIIR:Ag with the antiserum and staphylococcus protein A linked with horse-raddish peroxidase introduced by our laboratory confirmed its localization in endothelium cells, similar to the results obtained by classic indirect immunofluorescent staining. Three medical centers have been studying the carriers of hemophilia A. The results are shown in Table 3. It appeared that VIIIC/VIIIR:Ag was the most sensitive and simplest parameter for discriminating hemophilia A carriers. VIII:C/IX:C did not elevate the sensitivity of the discrimination.

Immunoluminometric assays for lactoferrin and elastase-alpha 1-proteinase inhibitor complexes were developed using solid-phase methodology, which has already been published from this laboratory. The aim of the study was to develop a rapid method to see whether elevated granulocyte activity was present in the lung, as for example in neonatal sepsis. The lactoferrin assay gave reliable results within 30 minutes, the elastase-alpha 1-proteinase inhibitor complexes, within 5 hours. The correlation between both analytes was good, so that the lactoferrin assay could replace the elastase-alpha 1-proteinase inhibitor assay in emergency cases. The lactoferrin assay was used for rapid answer, the elastase-alpha 1-proteinase inhibitor complex assay for "fine" monitoring of the progress of the disease. Both assays could be used to measure concentrations in plasma or bronchoalveolar lavage using a 10 microliters sample. Plasma for the elastase-alpha 1-proteinase inhibitor complex determination had to be diluted 1:50 before being assayed. Only EDTA plasma was used in the assay, as either heparin plasma or serum resulted in granulocyte destruction, thus giving rise to elevated, and non-reproducible results. The results from bronchoalveolar lavage show an excellent correlation between elastase-alpha 1-proteinase inhibitor complexes and lactoferrin. No interference was seen from lipaemic or icteric plasma samples. Results from haemolytic samples i.e. where lysis of erythrocytes and leukocytes had occurred, had to be treated with care if no clinical indication of intravascular haemorrhage was present. The assays lend themselves to perinatal diagnosis, as the total volume of plasma or lavage needed is theoretically under 50 microliters, i.e. ethically acceptable for regular monitoring of neonates.(ABSTRACT TRUNCATED AT 250 WORDS)

Abstract The genetic and biochemical basis of fish-eye disease (FED) was investigated in a 63-year-old female proband with low plasma HDL cholesterol. Analyses of corneal and plasma lipids of the proband were consistent with impaired lecithin:cholesterol acyltransferase (LCAT) activity. Free cholesterol and phospholipid levels were elevated relative to control values, whereas cholesteryl ester levels were greatly reduced. Fatty acid compositions of corneal lipids from the proband and control subjects differ from the respective fatty acid compositions of their plasma lipids. This suggests that the metabolic pathways and acyl chain specificities for phospholipid, cholesteryl ester, and triglyceride metabolism within the cornea are distinct from those of plasma. Sequencing of the LCAT gene from the proband revealed a novel mutation at nucleotide 399, corresponding to an Arg 99 →Cys substitution. Secretion of LCAT (Arg 99 →Cys) by transfected COS-6 cells was ≈50% of that of the wild type, but its specific activity against reassembled HDL was 93% lower than that of wild-type LCAT. The specific activities of wild-type and LCAT (Arg 99 →Cys) against LDL were reduced similarly, suggesting that the appearance of the FED phenotype does not require enhanced activity against LDL. Our data support the hypothesis that FED is a partial LCAT deficiency in which poor esterification in specific types of HDL particles may contribute to the appearance of the corneal opacities.

Giant cell angioblastoma (GCAB) is an extremely rare soft tissue tumor of early childhood and only five cases have been described to date. As such the clinical, pathological, and prognostic features are poorly defined. We prensent here a new case of GCAB in bone of a child aged 4-years old. The lesion was composed of dense and loose cell regions. The dense regions were characterized by nodular, linear, and plexiform aggregates of oval- to spindle-shaped tumor cells around small vascular channels and interspersed with large mononuclear cells and multinucleate giant cells. The loose cell areas were characterized by distributed fibroblasts and abundant myxoid matrix, which diminished with patient age. Infiltrative growth was observed in some areas. Oval-to-spindle cells showed positivity for Vimentin, CD31 and CD34 staining, and partial positivity for smooth muscle actin. Mononuclear cells and multinucleate giant cells showed Vimentin and CD68 positivity. Seventeen months after thorough curettage of the lesion, a local recurrence was found. Based upon the clinical, histological and immunohistochemical findings, infiltrate condition, and prognosis, we classified GCAB into two subtypes. Type I does not infiltrate surrounding tissues and has good prognosis. Type II infiltrates the surrounding tissues, relapses earlier, and has worse prognosis. This report augments the limited GCAB literature to promote our understanding and guide therapy of this rare disease. Virtual slides The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/6699811297488137

Compared to PET & MRI, there is a paucity of literature on normal myocardial blood flow (MBF) values measured by dynamic CT. Our aim was to 1) describe rest & stress MBF values in a healthy population using 128-slice dual source CT myocardial perfusion imaging (CTMPI), 2) compare MBF values of the

Li-Ting Chen,1,* Xiao-Le Fan,2,* Hai-Jun Li,1 Cheng-Long Ye,1 Hong-Hui Yu,1 Hui-Zhen Xin,1 Hong-Han Gong,1 De-Chang Peng,1 Li-Ping Yan3 1Department of Radiology, the First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi Province, China; 2Department of General Surgery, the First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi Province, China; 3Department of Cardiology, People’s Hospital of Jiangxi Province, Nanchang, Jiangxi Province, China *These authors contributed equally to this work Objective: Obstructive sleep apnea (OSA) is accompanied by widespread abnormal spontaneous regional activity related to cognitive deficits. However, little is known about the topological properties of the functional brain connectome of patients with OSA. This study aimed to use the graph theory approaches to investigate the topological properties and functional connectivity (FC) of the functional connectome in patients with OSA, based on resting-state functional magnetic resonance imaging (rs-fMRI). Methods: Forty-five male patients with newly diagnosed untreated severe OSA and 45 male good sleepers (GSs) underwent a polysomnography (PSG), clinical evaluations, and rs-fMRI scans. The automated anatomical labeling (AAL) atlas was used to construct the functional brain connectome. The topological organization and FC of brain functional networks in patients with OSA were characterized using graph theory methods and investigated the relationship between functional network topology and clinical variables. Results: Both the patients with OSA and the GSs exhibited high-efficiency “small-world” network attributes. However, the patients with OSA exhibited decreased σ, γ, Eglob; increased Lp, λ; and abnormal nodal centralities in several default-mode network (DMN), salience network (SN), and central executive network (CEN) regions. However, the patients with OSA exhibited abnormal functional connections between the DMN, SN, and CEN. The disrupted FC was significantly positive correlations with the global network metrics γ and σ. The global network metrics were significantly correlated with the Epworth Sleepiness Scale (ESS) score, Montreal Cognitive Assessment (MoCA) score, and oxygen desaturation index. Conclusion: The findings suggest that the functional connectome of patients with OSA exhibited disrupted functional integration and segregation, and functional disconnections of the DMN, SN, and CEN. The aberrant topological attributes may be associated with disrupted FC and cognitive functions. These topological abnormalities and disconnections might be potential biomarkers of cognitive impairments in patients with OSA. Keywords: obstructive sleep apnea, graph theory, functional connectome, cognitive function, resting-state fMRI

// Dong-dong Cheng 1, * , Hui-zhen Zhang 2, * , Jun-qing Yuan 2, * , Shi-jie Li 1 , Qing-cheng Yang 1 , Cun-yi Fan 1 1 Department of Orthopedics, Shanghai Jiao Tong University Affiliated Sixth People’s Hospital, Shanghai, 200233, China 2 Department of Pathology, Shanghai Jiao Tong University Affiliated Sixth People’s Hospital, Shanghai, 200233, China * Authors share co-first authorship Correspondence to: Cun-yi Fan, email: fancunyi888@hotmail.com Qing-cheng Yang, email: tjyqc@163.com Keywords: MCM2, MCM3, DHX9, proliferation, osteosarcoma Received: November 14, 2016 Accepted: February 07, 2017 Published: February 18, 2017 ABSTRACT A label free quantitative proteomic approach (SWATH™ experiment) was performed to identify tumor-associated nuclear proteins that are differentially expressed between osteosarcoma cells and osteoblast cells. By functional screening, minichromosome maintenance protein 2 (MCM2) and minichromosome maintenance protein 3 (MCM3) were found to be related to osteosarcoma cell growth. Here, we show that knockdown of MCM2 or MCM3 inhibits osteosarcoma growth in vitro and in vivo . In co-immunoprecipitation and co-localization experiments, MCM2 and MCM3 were found to interact with DExH-box helicase 9 (DHX9) in osteosarcoma cells. A rescue study showed that the decreased growth of osteosarcoma cells by MCM2 or MCM3 knockdown was reversed by DHX9 overexpression, indicating that MCM2 and MCM3 activity was DHX9-dependent. In addition, the depletion of DHX9 hindered osteosarcoma cell proliferation. Notably, MCM2 and MCM3 expression levels were positively correlated with the DHX9 expression level in tumor samples and were associated with a poor prognosis in patients with osteosarcoma. Taken together, these results suggest that the MCM2/MCM3–DHX9 axis has an important role in osteosarcoma progression.

// Qing-yong Chen 1, * , De-min Jiao 1, * , Yu-quan Wu 2 , Jun Chen 1 , Jian Wang 1 , Xia-li Tang 1 , Hao Mou 1 , Hui-zhen Hu 1 , Jia Song 1 , Jie Yan 1 , Li-jun Wu 1 , Jianyan Chen 3 , Zhiwei Wang 4, 5 1 Department of Respiratory Disease, The 117 th Hospital of PLA, Zhejiang, China 2 Department of Oncology, The 117 th Hospital of PLA, Zhejiang, China 3 Department of Anesthesiology, Shenzhen Baoan Hospital Affiliated to Southern Medical University, Guangdong, China 4 Department of Pathology, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, USA 5 The Cyrus Tang Hematology Center and Collaborative Innovation Center of Hematology, Jiangsu Institute of Hematology, The First Affiliated Hospital, Soochow University, Suzhou, China * These authors contribute equally to this work Correspondence to: Jianyan Chen, e-mail: chenjianyan@yahoo.com Zhiwei Wang, e-mail: zwang6@bidmc.harvard.edu Qing-yong Chen, e-mail: cqyong117@163.com Keywords: miR-206, HGF, epithelial-mesenchymal transition, angiogenesis, lung cancer Received: November 05, 2015 Accepted: February 9, 2016 Published: February 22, 2016 ABSTRACT MiR-206 is low expression in lung cancers and associated with cancer metastasis. However, the roles of miR-206 in epithelial-mesenchymal transition (EMT) and angiogenesis in lung cancer remain unknown. In this study, we find that hepatocyte growth factor (HGF) induces EMT, invasion and migration in A549 and 95D lung cancer cells, and these processes could be markedly inhibited by miR-206 overexpression. Moreover, we demonstrate that miR-206 directly targets c-Met and inhibits its downstream PI3k/Akt/mTOR signaling pathway. In contrast, miR-206 inhibitors promote the expression of c-Met and activate the PI3k/Akt/mTOR signaling, and this effect could be attenuated by the PI3K inhibitor. Moreover, c-Met overexpression assay further confirms the significant inhibitory effect of miR-206 on HGF-induced EMT, cell migration and invasion. Notably, we also find that miR-206 effectively inhibits HGF-induced tube formation and migration of human umbilical vein endothelial cells (HUVECs), and the mechanism is also related to inhibition of PI3k/Akt/mTOR signaling. Finally, we reveal the inhibitory effect of miR-206 on EMT and angiogenesis in xenograft tumor mice model. Taken together, miR-206 inhibits HGF-induced EMT and angiogenesis in lung cancer by suppressing c-Met/PI3k/Akt/mTOR signaling. Therefore, miR-206 might be a potential target for the therapeutic strategy against EMT and angiogenesis of lung cancer.