Cell Journal (Yakhteh)
Year:2009 | Volume:10 | Issue:4 (40)|Papers Count:8

Although the genome is defined by its primary sequence, its functional properties are determined by far more complex mechanisms and depend on multiple layers of nuclear organization. The architecture of the nucleus includes two overlapping structures: the chromatin and a framework structure named the nuclear matrix. Ultra-structural studies reveal that the nuclear matrix is a network consisting of branched core filaments masked with a large number of hnRNPs and regulatory proteins. This scaffold has been demonstrated to be an active and dynamic structure, anchoring the nuclear processes such as replication, transcription and splicing making nuclear domains/foci. It is postulated that the nuclear matrix serves as a dynamic support to bring together specific DNA sequences with factors involved in the regulation of genome functions. In this review, we attempt to introduce the structure and function of nuclear matrix as an active intra-nuclear factor, having a critical dynamic role to organize different nuclear functions. Studying in vivo variations of this epigenetic parameter has been suggested to all investigators interested in the field of chromatin structure and itsdynamics.

Objective: The role of Wnt signaling and its antagonist; secreted Frizzled Related Protein type 4 (sFPR4) was reported in rodent ovarian follicular development. This study examines immunolocalization of sFRP4 in ovaries of polycystic ovary (PCO) rat model and evaluates its role in follicular growth arrest and its premature differentiation.Materials and Methods: PCO was induced with daily administration of testosterone propionate (TP) for 1 to 4 weeks while normal control rats were injected only with vehicle. The ovaries underwent histological examination, immunohistochemical analysis of sFRP4 and steroidogenic acute regulatory protein (StAR) and apoptosis analysis.  Results: Four-week TP treatment significantly increased the primordial follicles, and significantly decreased the preantral and antral follicles compared to one week TP treatment. TP-treated animals had concomitantly, significant increase of sFPR4 immunoexpression in primordial, primary and preantral follicles as compared to one week TP-treated animals and control groups. Furthermore, sFRP4 immunostaining strongly co-localized in apoptotic granulosa cells. Interestingly, increased sFRP4 immunostaining was associated with increased StAR immunoexpression in follicular theca layer and stroma in four weeks TP-treated rats compared to one week TP-treated rats and control groups.Conclusion: Our data showed a highly significant association between sFRP4 expression and apoptosis in ovaries of four week TP-treated animals. Moreover, co-localization of StAR and sFRP4 could suggest that sFRP4 may play a role in premature differentiation of follicles. 

Objective: Evaluation of extracellular matrics (ECMs) effect on differentiation of embryonic stem cells (ESCs) to pancreatic β-cell.Materials and Methods: Mouse ESC line, Royan B1, was subjected to differentiation into b -like cells in a three-step method: generation of embryoid bodies (EBs), spontaneous differentiation and induction by Nicotinamide onto different matrices including poly L-ornithine/laminin, gelatin, and two different dilution of matrigel (1:30, 1:100) and control group (no ECM). At the final step, differentiated cells were analyzed for expression of some pancreas-specific genes using "semi-quantitative RT-PCR ", for detection of insulin and C-peptide presence in cells using "immunocytochemistry" and for the evaluation of the amount of secreted insulin in response to glucose Using "insulin secretion assay".Results: The semi-quantitative RT-PCR analysis of differentiated cells on 1:30 matrigel coated-plates showed consistent higher expression of b-cell specific markers including Insulin I, Insulin II, Slc2a2 in comparison to the other ECMs. The results of immunostainig for C-peptide showed no significant differences between the experimental groups and finally insulin secretion assay revealed that differentiated cells on 1:30 matrigel coated-plates secreted more insulin in response to glucose in comparison to the other ECMs.Conclusion: Our results suggest that type of ECM may influence ESC differentiation into insulin-secreting cells and 1:30 matrigel was more effective. However, the success rate of differentiation needs further investigations using other appropriate ECMs.

Objective: The purpose of the study was to evaluate the production of antibodies in serum as well as egg yolk raised against S. typhimurium and the cross-reactivity of this antibody with other enteric bacteria.Materials and Methods: White egg–laying hens were immunized with S. typhimurium, heat-killed whole cell, in Freund's adjuvant. Immunization was done with 107 conlony forming unit (CFU) of bacteria per hen which was injected into the breast muscle of lay. Specific antibodies were detected by enzyme-linked immunosorbent assay in the serum and in eggs. The serum and eggs of two adults white Leghorn hens were not immunized with S. typhimurium used as a control.Results: Chicken egg yolk antibodies (IgY) were raised against S. typhimuruim in the serum as well as in eggs. The production of IgY in serum was higher than IgY produced in egg yolk. Anti-S. typhimurium IgY cross-reacted 63%, 25% and 14% with S. typhimurium, Shigella dysenteriae  and E. coli respectively.Conclusion: The findings indicate that eggs from hens immunized with S. typhimuruim have not specific antibodies for the detection of S. typhimuruim, but they may have the potential of being a useful source of passive immunity against this pathogen.

Objective: The aim of this study was to select the best medium to maintain sperm motility during sperm-DNA incubation and assess the DNA uptake by spermatozoa of Iranian Holstein bulls and its effects on sperm motility.Materials and Methods: frozen sperms from an Iranian Holstein bull were thawed and centrifuged. Motile sperms were separated through Pure sperm gradient (40/80%) followed by two times washing in SP-TALP medium. Then, sperms were washed once (PBS, Opti-MEM and SP-TALP) and incubated with DNA in each media followed by sperm motility estimation. The plasmid pEGFP-C1 was linearized and incubated with sperms at 37°C for 1 hour. Sperm-DNA mixture was treated with DNase I and the sperm pellet was washed with PBS. DNA extraction from sperms and supernatants from the last washing were used as template for PCR. Data was analyzed using SAS package and mean comparisons between sperm motility in different media were performed.Results: Sperm motility after incubation in PBS, Opti-MEM and SP-TALP were 40(±2.89), 2(±1.53) and 54(±4.41) percent, respectively. PCR results from transfected sperms indicated that EGFP transgene internalized into the bovine sperms and DNaseI treatment could not eliminate it.Conclusion: In conclusion the best medium for sperm and DNA incubation was SPTALP. The DNA not only could attach to the post acrosomal region of spermatozoa but also could integrate into it. So bovine spermatozoa can be used as transgene carrier into oocyte.

Objective: To determine the role of global genome methylation in gastritis lesion and its relation with clinicopathologic finding.Materials and Methods: The study was conducted on 44 gastritis and normal adjacent specimens using a technique composed of restriction enzyme digestion and pyrosequencing known as LUMA (LUminometric Methylation Assay). At first, DNA extracted from gastritis lesion and normal tissue was digested with HpaII (sensitive to methylation in recognition site) and MspI (insensitive). These enzymes leave an overhang after cutting which are then filled in a polymerase extension assay with stepwise addition of dNTPs using pyrosequencing. The comparison of the height of picks obtained from both enzymes provides the possibility to evaluate and compare global genome methylation level of normal and gastritis tissues. If the target site is fully methylated, the HpaII /MspI will approach toward zero .If not, this ratio will go around one. In the other conditions the ratio varies between 0-1.Results: According to our findings, gastritis tissue was significantly more hypomethylated (p=0.04) than the nornal tissue and Global genome methylation had no correlation with sex, age, microsatellite instability (MSI) and gastritis severity. Conclusion: Global DNA hypomethylation occurs in the gastritis lesion. Presumably the process of hypomethylation keeps falling in the next steps leading to gastric cancer.

Objective: The urinary system of euryhaline fish, as well as the gills, is involved in ion regulation through the production of dilute urine in freshwater and isotonic urine in seawater. The low osmolality of the urine originates from an active reabsorption of ions by Ionocytes or mitochondria rich-cells (MRCs) present in certain parts of the urinary system. Mitochondria rich-cells possess a high density of Na+, K+-ATPase. Materials and Methods: Persian Sturgeon fry’s, adapted to freshwater and diluted Caspian Sea water (‰5 salinity) were fixed in Bouin’s solution after 24h. After the hydration with ethanol, the samples were paraffinaized and sectioned. Light microscopy and Hematoxiline-Fushin staining were used for histological examinations. Immunolocalization of Na+, K+-ATPase was observed through fluorescent microscopy (450-490mm), using IgGa5 (as primary antibody) and FITC (as secondary antibody).Results: In both experimental conditions, maximum immunofluorescence of Na+, K+-ATPase (in mitochondria rich-cells) was found in distal and collective tubules. In both ureter and urinary bladder, immunostainings were found in dispersed cells with relatively weak intensity. In ‰5 acclimated fish, weak immunofluorescence was also observed in neck segment and proximal tubules, as well as in distal and collective tubules.Conclusion: Alternation in mitochondria rich-cells and Na+, K+-ATPase distribution in kidney tubules of ‰5 acclimated fry’s showed that the blood and osmolytes were nearly isotonic to environment but not isoionic. Thus the fish needs the absorption and excretion of some ions for the body homeostasis and osmoregulation.

Objective: The present retrospective study aims at identifying the prevalence of chromosomal abnormalities in a population of couples who are candidates for assisted reproductive techniques.Materials and Methods: Cytogenetic analysis was performed according to the standard methods on cultured cells from the patients’ peripheral blood. The culture was harvested after 72 hours. At least 20 metaphases were examined by GTG banding.Results: The analyses of the Karyotypes of 1726 candidate patients (863 men and 863 women) revealed a total of 107 aberrant karyotypes, the frequencies of abnormalities were 3.6% (31/863) for men and 8.8% (76/863) for women. The following frequencies of abnormalities were observed for women: 6% (n=52) for sex chromosome mosaicism, 1.4% (n=12) for translocations, 0.3% (n=3) for inversions and 1% (n=9) for other abnormalities. Whereas, the frequencies of abnormalities for men were: 0.9% (n=8) for sex chromosome mosaicism, 1.2% (n=11) for translocations, 0.3% (n=3) for autosomal inversions, 0.4% (n=4) for Y chromosome inversion. Kelinefelter syndrome was detected in one patient and 47, XY, +mar karyotype in another patient.Conclusion: The high frequency of chromosomal abnormalities in couples with reproductive failure is well distinguished. In this study a higher frequency of aberrations was more observed in women than men. The frequency of translocations was similar in both sexes, but in men the frequency of inversions was higher than women, whereas the frequency of sex chromosome mosaicism in women was higher than men.