Whole-genome methylation analysis reveals epigenetic variation between wild-type and nontransgenic cloned,ASMT transgenic cloned dairy goats generated by the somatic cell nuclear transfer

Background:SCNT(somatic cell nuclear transfer)is of great significance to biological research and also to the livestock breeding.However,the survival rate of the SCNT cloned animals is relatively low compared to other transgenic methods.This indicates the potential epigenetic variations between them.DNA methylation is a key marker of mammalian epigenetics and its alterations will lead to phenotypic differences.In this study,ASMT(acetylserotonin-Omethyltransferase)ovarian overexpression transgenic goat was produced by using SCNT.To investigate whether there are epigenetic differences between cloned and WT(wild type)goats,WGBS(whole-genome bisulfite sequencing)was used to measure the whole-genome methylation of these animals.Results:It is observed that the different m Cp G sites are mainly present in the intergenic and intronic regions between cloned and WT animals,and their CG-type methylation sites are strongly correlated.DMR(differentially methylated region)lengths are located around 1000 bp,mainly distributed in the exonic,intergenic and intronic functional domains.A total of 56 and 36 DMGs(differentially methylated genes)were identified by GO and KEGG databases,respectively.Functional annotation showed that DMGs were enriched in biological-process,cellularcomponent,molecular-function and other signaling pathways.A total of 10 identical genes related to growth and development were identified in GO and KEGG databases.Conclusion:The differences in methylation genes among the tested animals have been identified.A total of 10 DMGs associated with growth and development were identified between cloned and WT animals.The results indicate that the differential patterns of DNA methylation between the cloned and WT goats are probably caused by the SCNT.These novel observations will help us to further identify the unveiled mechanisms of somatic cell cloning technology,particularly in goats.

Proliferating cell nuclear antigen clamp associated factor,a potential proto-oncogene with increased expression in malignant gastrointestinal tumors

Gastrointestinal(GI)cancers,including malignancies in the gastrointestinal tract and accessory organs of digestion,represent the leading cause of death worldwide due to the poor prognosis of most GI cancers.An investigation into the potential molecular targets of prediction,diagnosis,prognosis,and therapy in GI cancers is urgently required.Proliferating cell nuclear antigen(PCNA)clamp associated factor(PCLAF),which plays an essential role in cell proliferation,apoptosis,and cell cycle regulation by binding to PCNA,is a potential molecular target of GI cancers as it contributes to a series of malignant properties,including tumorigenesis,epithelial-mesenchymal transition,migration,and invasion.Furthermore,PCLAF is an underlying plasma prediction target in colorectal cancer and liver cancer.In addition to GI cancers,PCLAF is also involved in other types of cancers and autoimmune diseases.Several pivotal pathways,including the Rb/E2F pathway,NF-κB pathway,and p53-p21 cascade,are implicated in PCLAF-mediated diseases.PCLAF also contributes to some diseases through dysregulation of the p53 pathway,WNT signal pathway,MEK/ERK pathway,and PI3K/AKT/mTOR signal cascade.This review mainly describes in detail the role of PCLAF in physiological status and GI cancers.The signaling pathways involved in PCLAF are also summarized.Suppression of the interaction of PCLAF/PCNA or the expression of PCLAF might be potential biological therapeutic strategies for GI cancers.

Expression of germ cell nuclear factor in mouse germ cells and sperm during postnatal period

Aim: To assess the spatial and temporal expression of germ cell nuclear factor (GCNF) in male mouse germ cells during postnatal development and in sperm before and after capacitation. Methods: The indirect immun-ofluorescence method with anti-GCNF antiserum was used to investigate the GCNF expression in mice at day 8, 10, 14, 17, 20, 28, 35, 70, and 420 after birth and in sperm before and after capacitation. Results: With the proceeding of spermatogenesis, GCNF was first detected in the nuclei of spermatogonia and a few early stage primary sperma-tocytes at day 8, which was increased gradually at day 10 to 14 inclusive. From day 17 to day 20, the GCNF was concentrated in round spermatids, while both spermatogonia and early stage primary spermatocytes became GCNF negative. From day 28 until day 420, strong GCNF expression was shown in round spermatids and pachytene spermatocytes, while spermatogonia, early primary spermatocytes and elongating spermatids were all GCNF negative. In addition, it was also found that GCNF was localized on the acrosomal cap region of spermatozoa and there was a big change in GCNF expression during capacitation, from 98 % GCNF positive before capacitation to about 20 % positive following capacitation. The localization of GCNF in caput and cauda spermatozoa was similar. Conclusion: GCNF may play important roles in spermatogenesis, capacitation and fertilization.

Evaluation of germ-cell kinetics in infertile patients with proliferating cell nuclear antigen proliferating index

Aim: To explore the usefulness of proliferating cell nuclear antigen proliferating index (PCNA PI) in the pathologicaldiagnosis and treatment of male infertility. Methods: Testicular biopsy specimen obtained from 48 cases of male in-fertility and 2 normal controls were fixed and embedded. The sections were stained with anti-PCNA monoclonal anti-bodies or haematoxylin/eosin. Proliferating index (PI), expressed as the percentage of germ-cell nuclei positivelystained with PCNA antibody, was assessed from more than 20 seminiferous tubules or 600 germ-cells. Results: Theinfertile patients were divided into 4 groups: Group 1, normal spermatogenesis (14 cases); Group 2, hypospermato-genesis (16 cases); Group 3, germinal arrest (10 cases); Group 4, Sertoli cell only syndrome (8 cases). The PCNAPI of normal control testis was 86.5 % (mean value). Group 3 had a significantly lower PCNA PI (29.8 %) than nor-mal testis; Group 1 and 2 had similar Pis (82.3% and 82.3%, respectively) as the control testis. PI of the negativecontrol (Group 4) was 0 as no germ-cells were found. Conclusion: PCNA PI is useful for assessing germ-cell ki-netics, especially for pathological diagnosis of germinal arrest which is difficult to differentiate by routine HE stainingtechnique. In germinal arrest, there is a significantly lowered PCNA PI, which is an indication of DNA synthesis dete-rioration, suggesting the use of therapies be different from those for hypospermatogenesis. (Asian J Androl 2001 Mar;3: 63-66)

The Relationship of Expression of bcl-2, p53, and Proliferating Cell Nuclear Antigen (PCNA) to Cell Proliferation and Apoptosis in Renal Cell Carcinoma

To investigate the relationship of bcl-2, p53, proliferating cell nuclear antigen (PCNA) to cell proliferation, apoptosis and pathological parameters, the patterns of cell growth and turnover in renal cell carcinoma (RCC), formalin-fixed and paraffin-embedded tissue blocks from 34 patients with RCC were examined. Cell proliferation activity was detected by PCNA immunostaining and the proliferation index (PI) was expressed as a percentage of the PCNA-positive cells in the tumor cells. Apoptosis was detected by terminal deoxy- nucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL), and the apoptotic index (AI) was expressed as a percentage of the TUNEL-positive cells in the tumor cells. Expressions of bcl-2 and p53 were assessed immunohistochemically. Our results showed that the PI ranged from 6.0 % to 24.0 % (median 12.3 %) and the AI from 2.0 % to 8.0 % (median 5.4 %) in RCC. The expression of the bcl-2 protein was demonstrated in 15 cases (44.1 %); the expression of the p53 protein, however, was seen in only 3 case. bcl-2 positivity was not associated with PI or AI or any pathological parameters. There were close associations between PI and tumor grade and stage, and a significant relationship between AI and the tumor grade of RCC. Our study suggests that bcl-2 positivity was not associated with PI or AI or any pathological parameters. There are close associations between PI and AI and tumor grade and stage of RCC. Active cell proliferation may be accompanied by frequent apoptosis in RCC.

Spatial and temporal expression of germ cell nuclear factorin murine epididymis

Aim: To investigate the spatial and temporal expression of germ cell nuclear factor (GCNF) in mouse and rat epididymis during postnatal period. Methods: The epididymal sections from different postnatal days were stained for GCNF by the indirect immunofluorescence technique and digital photographs were taken by a Carl Zeiss confocal microscope. Results: GCNF was first detected on day 12 in mouse epididymis and day 14 in rat epididymis. The highest expression of GCNF was observed on day 35 in both mouse and rat epididymis. In adults, GCNF exhibited a region-specific expression pattern, i.e., it was expressed predominantly in the initial segment, caput and proximal corpus of rat epididymis and was abundant in the proximal corpus of mouse epididymis. GCNF could be found in the nuclei of the principal, apical, narrow, clear and halo cells. Conclusion: GCNF may play an important role in epididymal differentiation and development and in sperm maturation.

Influence of norcantharidin on proliferation,proliferation-related gene proteins prolifera-ting cell nuclear antigen and Ki-67 of human gallbladder carcinoma GBC-SD cells

BACKGROUND: Gallbladder carcinoma is a highly lethal and aggressive disease with early metastasis, strong invasion and poor prognosis. Most patients with this disease are at the advanced and un-resectable stage and should be consi- dered for palliative treatment such as chemotherapy and ra- diotherapy. Unfortunately, reports of chemotherapy and radiotherapy for gallbladder carcinoma are disappointing. We investigated the influence of norcantharidin (NCTD) on proliferation, proliferation-related gene proteins PCNA and Ki-67 of human gallbladder carcinoma GBC-SD cells in vitro. METHODS: GBC-SD cell lines of human gallbladder carci- noma were cultured by the cell culture technique. The ex- periment was divided into NCTD group and control group. The tetrazolium-based colorimetric assay was used to evaluate cell growth. The streptavidin-biotin complex method was used to determine the expressions of prolifera- tion-related gene proteins PCNA and Ki-67 of human gall- bladder carcinoma GBC-SD cells. RESULTS: NCTD inhibited the growth and proliferation of GBC-SD cells from 10 mg/L or after 6 hours in a dose- and time-dependent manner, with the IC50 value of 56.18 μg/ ml at 48 hours. After treatment with NCTD, the expression of PCNA (0.932 ±0.031 vs. 0.318 ±0.023, P<0.001) and Ki-67 (0.964 ±0.092 vs. 0.297 ±0.018, P<0.001) proteins were decreased significantly. CONCLUSION: NCTD inhibits the proliferation of human gallbladder carcinoma GBC-SD cells in vitro and the expres- sion of their proliferation-related gene proteins PCNA and Ki-67.

The Relationship between Apoptosis and the Expression of Proliferating Cell Nuclear Antigen and the Clinical Stages in Gastric Carcinoma

The relationship between the apoptosis and the expression of proliferating cell nuclear antigen (PCNA) and the clinical stages in gastric cancers was studied. By using terminal deoxynucleotidyl transferase mediated nick end labelling (TUNEL) technique and PCNA immunohistochemical staining, the apoptosis and the expression of PCNA in tissue of gastric carcinoma were assayed in situ, the index of apoptosis (AI), index of PCNA (PI) and the rate of AI/PI were calculated. AI and PI in gastric cancer tissues were (6.5±3.7) % and (49.8±15.9) % respectively, and the rate of AI/PI was 0.13±0.05, which were obviously different from those of normal gastric mucosa in paragastric cancer ( P <0.01). With the advanced TNM stages of gastric carcinoma, the AI was decreased, PI was increased and the rate of AI/PI decreased in gastric carcinoma. There was significant difference in them between the gastric cancer tissues and normal gastric mucosa in pericarcinoma in TNM stage Ⅱ to Ⅳ ( P <0.05). It was suggested that the decreased apoptotic cells and the increased proliferating cells were obviously related to the tumor genesis and tumor progression in gastric carcinoma. The AI, PI and the rate of AI/PI would become the prognostic factors in advanced gastric carcinoma.

RELATIONSHIP BETWEEN PROLIFERATING CELL NUCLEARANTIGEN EXPRESSION AND ITS MALIGNANCY POTENTIAL IN COLORECTAL CARCINOMA

Objective: To study the relationship between proliferating cell nuclear antigen expression and its malignancy potential in colorectal carcinoma. Methods: Paraffin sections of 86 patients with advanced colorectal carcinoma were assessed by immunohistochemical study, using a mouse monoclonal antibody (pc-10, DAKO Co. USA) to check proliferating cell nuclear antigen (PCNA). To compare PCNA with conventional clinicopathologic factor, including p53 overexpression, tissue carcinoembnyonic antigen immunoreactivity pattern and flow cytometric DNA ploidy for assessing tumor malignancy potential. In addition, recurrence and survival of patients with advanced colorectal carcinoma after curative resection were analyzed in accordance with degree of PCNA expression. Results: PCNA-labeling index (PCNA-LI) increased significantly as the tumor stage advanced (p=0.0001). Strong correlations were observed between PCNA-LI and various pathologic parameters, including histologic differentiation (P=0.0027), lymphatic invasion (P=0.0001), vascular invasion (P=0.0001), lymph node metastasis (P=0.0001), and liver metastasis (P=0.0036). Mean PCNA-LI was also significantly higher in tumor with DNA aneuploidy (P=0.0006) and negative (P=0.01). Linear relationships were demonstrated between PCNA-LI and clinical outcomes; Recurrence rate was significantly greater in the group with higher than the mean PCNA-LI, who underwent curative resection (P<0.01), and three-year survival rates for curative cases with higher than the mean PCNA-LI were significantly poorer than those with lower than mean PCNA-LI (P<0.005). Conclusion: There were correlations between PCNA-LI and various pathologic parameters, PCNA-LI increased significantly as the tumor stage advanced in colorectal carcinoma, the rates of recurrence and death got higher as PCNA-LI increased after curative resection for colorectal carcinoma.

Cloning and characterization of proliferating cell nuclear antigen gene of Alexandrium catenella (Dinoflagellate) with respect to cell growth

Harmful algal blooms （HABs） have been affecting negatively the shellfish and aquaculture industries around the world. Though a lot of efforts have been made to disclose the changes of environmental factors involved and their effects on the HABs events, the molecular mechanism of this process remains unclear. To address this problem, proliferating cell nuclear antigen gene （pcna） was isolated and characterized from Alexandrium catenella. It showed high homology to those of other dinoflagellates （89% and 91% homology to Pfiesteria piscicid and Pyrocystis lunula, respectively）, and also 42%–43% homology to those of plant and animals. The expression level of pcna revealed by quantitative real time PCR was the lowest at the late lagging cell growth phase, increased to the highest at the late exponential phase, and then decreased at the stationary phase. Though the cell growth rate was also changing, no positive correlation between pcna expression level and cell growth rate was displayed throughout the whole cell growth stages （r 2 =0.024 6）. However, the pcna expression level had the similar trend with the change of cell growth rate throughout the whole growing process, e.g., from increasing at the earlier cell growth stage to decreasing at the following stages, though slightly lagging to the latter.

EXPRESSION OF P53 PROTEIN AND PROLIFERATING CELL NUCLEAR ANTIGEN IN HUMAN GESTATION TROPHOBLASTIC DISEASE

To study the relationship between p53 protein, proliferating cell nuclear antigen (PCNA) expression and benign or malignant gestational trophoblastic disease (MGTD). Methods: The histotomic sections of 48 patients with gestational trophoblastic disease and 24 patients of normal chorionic villi were stained using immunohistochemistry. The monoclonal antibodies were used to determine p53 protein and PCNA. Results: The frequency of p53 and PCNA positive expression were significantly different among the chorionic villi of normal pregnancy, hydratidiform mole (HM) and MGTD. But neither p53 nor PCNA has any relation with the clinical staging or metastasis of MGTD. Conclusion: Both P53 and PCNA are valuable in diagnosis of human gestational trophoblastic disease.

Abnormal expressions of proliferating cell nuclear antigen and P27 protein in brain glioma

Both proliferating cell nuclear antigen and P27 protein are important factors to regulate cell cycle. While, the combination of them can provide exactly objective markers to evaluate prognosis of patients with brain glioma needs to be further studied based on pathological level. OBJECTIVE： To observe the expressions of proliferating cell nuclear antigen and P27 protein in both injured and normal brain glioma tissues and analyze the effect of them on onset and development of brain glioma. DESIGN： Case contrast observation. SETTING： Department of Neurosurgery, the Second Affiliated Hospital of Xi＇an Jiaotong University. PARTICIPANTS： A total of 63 patients with brain glioma were selected from Department of Neurosurgery, the Second Affiliated Hospital of Xi＇an Jiaotong University from July 1996 to June 2000. There were 38 males and 25 females and their ages ranged from 23 to 71 years. Based on pathological classification and grading standards of brain glioma, patients were divided into grade I - II （n=30） and grade III- IV （n = 33）. All cases received one operation but no radiotherapy and chemiotherapy before operation. Sample tissues were collected from tumor parenchyma. Non-neoplastic brain tissues were collected from another 12 non-tumor subjects who received craniocerebral trauma infra-decompression and regarded as the control group. There were l0 males and 2 females and their ages ranged from 16 to 54 years. The experiment had got confirmed consent from local ethic committee and the collection was provided confirmed consent from patients and their relatives. All samples were restained with HE staining so as to diagnose as the brain glioma. While, all patients with brain glioma received radiotherapy after operation and their survival periods were followed up. METHODS： Primary lesion wax of brain glioma was cut into serial sections and stained with S-P immunohistochemical staining. Brown substance which was observed in tumor nucleus was regarded as the positive expressions of both proliferating cell nuclear antigen and P27 protein. Automatic imaging analytic system was used to quantitatively analyze staining results of tumor. MAIN OUTCOME MEASURES： To compare the expressions of proliferating cell nuclear antigen and P27 protein in brain glioma tissues and non-tumor brain tissues and investigate the effect of various sexes, ages, survival periods and severities on the expressions of them in brain tissues. RESULTS： There was no significant difference of sexes and ages in the expressions of proliferating cell nuclear antigen and P27 protein （P 〉 0.05）; however, the expressions of proliferating cell nuclear antigen and P27 protein were milder in non-tumor brain tissues than those in the brain glioma tissues （P 〈 0.05）. Expression of proliferating cell nuclear antigen in brain tissue of grade III- IV severity was stronger than that of grade I - II severity, and the expression in ≥ 5-year survival periods were also stronger than that in 〈 5-year survival periods （P 〈 0.05）. In addition, expression of P27 protein in brain tissue of grade III- IV severity was stronger than that of grade I - II severity, and the expression in ≥ 5-year survival periods were also stronger than that in 〈 5-year survival periods （P 〈 0.05）. CONCLUSION： Abnormal expressions of proliferating cell nuclear antigen and P27 protein in human brain glioma are closely related to onset, development and prognosis of tumor.

Neuron specific enolase,p53蛋白和proliferating-cell nuclear antigen在肺癌组织中的表达及意义

目的：研究神经元特异性烯醇化酶（neuronspecificenolase，NSE）的表达，与p53蛋白和增殖细胞核抗原（prolif－erating－cellnuclearantigen，PCNA）表达的关系及意义．方法：用特异性鼠抗人单克隆抗体，按LSAB免疫组织化学方法检测石蜡包埋的肺癌标本中NSE，p53蛋白和PCNA的表达．结果：在小细胞性肺癌（SmallCellLungCancer，SCLC）中，NSE阳性表达者，PCNA标记指数（LabellingIndex，LI）高于NSE阴性者，而p53蛋白的表达与NSE的表达无关．在非小细胞性肺癌（non－smallcelllungcancr，non－SCLC）中，p53蛋白的表达和PCNALI均与NSE的表达无关．结论：在SCLC的发生及发展过程中，神经内分泌功能可能起了部分作用．而p53抑癌基因在SCLC的发生中并不起着重要的作用．

Effects of UVB on the expression of proliferating cell nuclear antigen (PCNA) in dinoflagellate Prorocentrum donghaiense Lu

PCNA has been considered as a useful marker for the estimation of growth rates of marine phytoplankton at the species level. Since dinoflagellates are noted for having many prokaryotic features in that they are the only eukaryotes to have permanently condensed chromosomes as well as lacking histones and a nucleosome, the sensitivity to UVB radiation and the validity of PCNA as a maker of growth rate in dinoflagellate need to be evaluated. Prorocentrum donghaiense Lu was investigated to valuate the UVB sensitivity in relation to cellular and molecular aspects of PCNA as a growth indicator. The effects of UVB radiation on PCNA were studied using the methods of western blots technology and whole-cell immunoflurescence with one mono-antibody. UVB changed the cell morphology, halted the growth and increased the cell size, even caused cell death to a certain extent after treatment with UVB radiation in P. donghaiense. Compared with the control, treating the algal cultures in exponential phases with UVB radiation for 24 h caused chromatin release and increases in protein levels of PCNA.

The Effectof the Lyso PC-induced Endothelial Cell Conditioned Medi-um on Proliferating Cell Nuclear Antigen Expression of the Calf Tho-racic Aorta Smooth Muscle Cell

In order to study the effect of and mechanism of lysophosphatidylcholine (LysoPC) on proliferation of the calf thoracic aorta smooth muscle cells (ASMCs), the ASMCs were used to observe the effects of LysoPC induced endothelial cell conditioned medium on the DNA content and proliferating cell nuclear antigen (PCNA) expression in the calf thoracic ASMCs by flow cytometry and Western Blot technique. It was found that LysoPC induced endothelial cell conditioned medium could significantly promote PCNA expression of the calf ASMCs, induce the converting of ASMCs from G 0 /G 1 phase to S phase of DNA synthesis, and increase the tyrosine phosphorylation protein expression. Tyrosine protein kinase inhibitor (TPKi) RG50864 could obviously inhibit proliferation of LysoPC induced ASMCs in a dose dependence manner. The results indicated that the effect of LysoPC promoting the proliferation of ASMCs is partly evoked by endothelial cell derived growth factors such as PDGF and so on.

EXPRESSIONS OF P_(53), PROLIFERATING CELL NUCLEAR ANITIGEN,BCL-2 PROTEIN AND THEIR SIGNIFICANCE IN SALIVARY ADENOID CYSTIC CARCINOMA

Objective To study the effects of P53, PCNA, Bc1-2 protein and their relationship in salivary adenoid cystic carclnoma(SACC). Methods These protelns were examlned by lmmunohistochemistry. Results overexpressions of Ps, and PCNA were revealed in ACC samples, they were higher than those in (polymorphous adenomas) PA, but expression of Bc1-2 protein was not different between ACC and PA. In 3 subtypes of ACC, expressions of 3 proteins were different. Conciusion Mutations of P53, Bc1-2 may be involed in the occurrence of SACC, expression of PCNA and mutation of P53 may coexist in the development of the SACC.

Clinical significance of expression of proliferating cell nuclear antigen and E-cadherin in gastric carcinoma

AIM to investigate the expression of proliferating cell nuclear antigen(p CNA)and E-cadherin in gastric carcinoma and to analyze their clinical significance.METHODS A total of 146 patients were selected for this study,including 38 patients with intestinal metaplasia,42with dysplasia,and 66 with primary gastric cancer.In addition,40 patients with normal gastric tissues were selected as controls.the expression of p CNA and E-cadherin was detected by immunohistochemistry.Differences in p CNA and the E-cadherin labeling indexes among normal gastric mucosa,intestinal metaplasia,dysplasia,and gastric carcinoma were compared.Subjects with normal gastric tissues were assigned to a normal group,while gastric cancer patients were assigned to a gastric cancer group.the difference in p CNA and E-cadherin expression between these two groups was compared.the relationship between expression of p CNA and E-cadherin and clinicopathological features was also explored in gastric cancer patients.furthermore,prognosis-related factors,as well as the expression of p CNA and E-cadherin,were analyzed in patients with gastric cancer to determine the 3-year survival of these patients.RESULTS the difference in p CNA and the E-cadherin labeling indexes among normal gastric mucosa,intestinal metaplasia,dysplasia,and gastric carcinoma was statistically significant(p<0.05).During the transition of normal gastric mucosa to gastric cancer,the p CNA labeling index gradually increased,while the E-cadherin labeling index gradually decreased(p<0.05).the p CNA labeling index was significantly higher and the E-cadherin labeling index was significantly lower in gastric cancer than in dysplasia(p<0.05).the expression of p CNA was significantly higher in the gastric cancer group than in the normal group,but E-cadherin was weaker(p<0.05).there was a negative correlation between the expression of p CNA and E-cadherin in gastric carcinoma(r=-0.741,p=0.000).p CNA expression differed significantly between gastric cancer patients with and without lymph node metastasis and between patients at different t stages.E-cadherin expression also differed significantly between gastric cancer patients with and without lymph node metastasis(p<0.05).High t stage and positive p CNA expression were risk factors for the prognosis of patients with gastric cancer(RR>1),while the positive expression of E-cadherin was a protective factor(RR<1).the sensitivity,specificity,and accuracy of p CNA positivity in predicting the 3-year survival of patients with gastric cancer were 93.33%,38.89%,and0.64,respectively;while these values for E-cadherin negativity were 80.0%,41.67%,and 0.59,respectively.When p CNA positivity and E-cadherin negativity were combined,the sensitivity,specificity,and accuracy were66.67%,66.67%,and 0.67,respectively.CONCLUSION Combined detection of p CNA and E-cadherin can improve the accuracy of assessing the prognosis of patients with gastric cancer.

Mast cell density and the context of clinicopathological parameters and expression of p185,estrogen receptor,and proliferating cell nuclear antigen in gastric carcinoma

AIM: To investigate the relationship between the mast cell density (MCD) and the context of clinicopathological parameters and expression of p185, estrogen receptor (ER), and proliferating cell nuclear antigen (PCNA) in gastric carcinoma.METHODS: Mast cell, p185, ER, and PCNA were detected using immunohistochemical S-P labeling method. Mast cell was counted in tissue of gastric carcinoma and regional lymph nodes respectively, and involved lymph nodes (TLN) were examined as usual.RESULTS: MCD was significantly related to both age and depth of penetration (χ2=4.688,P＜0.05 for age and χ2=9.350,P＜0.01 for depth of penetration) between MCD＞21/0.03 mm2 and MCD≤21/0.03 mm2 in 100 patients; MCD in 1-6 ILN group patients was significantly higher than that in 7-15 TLN or ＞15 TLN group patients (u=6.881, 8.055, P＜0.01);There were significant differences intergroup in positive expression rate of p185, ER and PCNA between MCD ＞21/0.03 mm2 and MCD≤21/0.03 mm2 in 100 patients.CONCLUSION: Mast cell may have effect on inhibiting invasive growth of tumor, especially in the aged patients; The number of mast cells, in certain degree, may predicate the number of involved lymph nodes, which is valuable for assessment of prognosis; MCD was related to the expression of p185, ER, and PCNA in gastric carcinoma. Tt suggests that mast cell accumulation may inhibit the proliferation and the dissemination of the gastric carcinoma.

Cumulus-specific genes are transcriptionally silent following somatic cell nuclear transfer in a mouse model

This study investigated whether four cumulus-specific genes: follicular stimulating hormone receptor (FSHr), hyaluronan synthase 2 (Has2), prostaglandin synthase 2 (Ptgs2) and steroidogenic acute regulator protein (Star), were correctly reprogrammed to be transcriptionally silent following somatic cell nuclear transfer (SCNT) in a murine model. Cumulus cells of C57×CBA F1 female mouse were injected into enucleated oocytes, followed by activation in 10 μmol/L strontium chloride for 5 h and subsequent in vitro culture up to the blastocyst stage. Expression of cumulus-specific genes in SCNT-derived embryos at 2-cell, 4-cell and day 4.5 blastocyst stages was compared with corresponding in vivo fertilized embryos by real-time PCR. It was demonstrated that immediately after the first cell cycle, SCNT-derived 2-cell stage embryos did not express all four cumulus-specific genes, which continually remained silent at the 4-cell and blastocyst stages. It is therefore concluded that all four cumulus-specific genes were correctly reprogrammed to be silent following nuclear transfer with cumulus donor cells in the mouse model. This would imply that the poor preimplantation developmental competence of SCNT embryos derived from cumulus cells is due to incomplete reprogramming of other embryonic genes, rather than cumulus-specific genes.

Production of homeobox A10 gene transgenic pigs by somatic cell nuclear transfer

Homeobox A10(Hoxa10) gene is one of the most important candidate genes associated with the reproductive performance of humans and mice. Overexpression of Hoxa10 in mouse endometrium can increase litter size. Moreover, Hoxa10 plays a key role in regulating the embryo implantation of sows. This study aimed to generate transgenic pigs using Hoxa10 via somatic cell nuclear transfer(SCNT). We established seven Hoxa10-transgenic cell lines, and two of the cell lines were selected as nuclear donors for the transfer. A total of 1 270 cloned embryos were generated and transferred to five surrogate mothers(Landrace×Yorkshire). Eight cloned male piglets were produced including one with cryptorchidism. Six transgenic piglets grew up healthy and produced 56 offspring. Finally, we obtained six transgenic male pigs and 26 transgenic positive offspring that can be used to further study the regulatory mechanism of Hoxa10 on the reproductive performance of pigs.