Calcium alginate microencapsulation of ovarian follicles impacts FSH delivery and follicle morphology

BACKGROUND: We have previously shown that suspension culture prevents follicle flattening and maintains three-dimensional follicle architecture better than culture on flat plates. However, many of the follicles cultured in suspension do eventually rupture, as basement membrane integrity is lost and the three-dimensional structure of the follicle is altered. Therefore, the objective of this study is to support three-dimensional follicle architecture during in vitro growth of ovarian follicles through encapsulation in calcium alginate, while maintaining responsiveness to FSH stimulation. METHODS: Preantral follicles (150 – 160 micrometers in diameter) were isolated from the ovaries of juvenile rats and grown in culture tubes or encapsulated in calcium alginate and grown in culture tubes. Previous studies revealed that follicles maintained structural integrity but did not grow as well when encapsulated in calcium alginate. In these studies, we evaluated the effect of calcium alginate on FSH-stimulated follicle growth, survival, and morphology in suspension culture. Follicles were grown under 5 culture conditions: 1) not encapsulated; with FSH in the medium, 2) encapsulated in the absence of FSH, grown in medium without FSH, 3) encapsulated with calcium alginate containing FSH but grown in medium without FSH, 4) encapsulated without FSH but grown in medium containing FSH and 5) encapsulated with calcium alginate containing FSH and in medium containing FSH. To assess growth rates, follicles were cultured for 72 hours and analyzed for follicle size increase and DNA content. Survival analysis for encapsulated and unencapsulated follicles was performed by constructing a Kaplan Meier survival curve of daily observations of intact follicle survival. Three-dimensional architecture was assessed histologically and by analysis of the pattern of connexin 43 expression in the cultured follicles. RESULTS: In the absence of FSH, follicle diameter increased by only 6.4%. When FSH was included in the alginate bead alone or the media alone, the follicle diameter increased by 13.5% and 19.9% respectively. This was greater than follicles cultured in the absence of FSH (p < 0.05), but less than that of the FSH-treated unencapsulated follicles (p < 0.05). However, when follicles were cultured with FSH included in both the media and the bead, a 32.6% increase in follicle diameter was observed, statistically no different than the growth rate of the unencapsulated follicles grown with FSH. CONCLUSION: Microencapsulation supports three-dimensional follicle growth, but may limit access to hormones in the medium resulting in altered development compared to unencapsulated follicles. Inclusion of FSH in the alginate bead restores the follicle growth response to FSH, while also providing a scaffold of support for three-dimensional growth. The application of tissue engineering principles to the problems of follicle culture in vitro may provide advances applicable to fertility preservation in women and endangered species

Interactions between the RepB initiator protein of plasmid pMV158 and two distant DNA regions within the origin of replication

Plasmids replicating by the rolling circle mode usually possess a single site for binding of the initiator protein at the origin of replication. The origin of pMV158 is different in that it possesses two distant binding regions for the initiator RepB. One region was located close to the site where RepB introduces the replication-initiating nick, within the nic locus; the other, the bind locus, is 84 bp downstream from the nick site. Binding of RepB to the bind locus was of higher affinity and stability than to the nic locus. Contacts of RepB with the bind and nic loci were determined through high-resolution footprinting. Upon binding of RepB, the DNA of the bind locus follows a winding path in its contact with the protein, resulting in local distortion and bending of the double-helix. On supercoiled DNA, simultaneous interaction of RepB with both loci favoured extrusion of the hairpin structure harbouring the nick site while causing a strong DNA distortion around the bind locus. This suggests interplay between the two RepB binding sites, which could facilitate loading of the initiator protein to the nic locus and the acquisition of the appropriate configuration of the supercoiled DNA substrate

The Effectiveness of Individually Guided Education on the Development of Self-Esteem in Middle School Students

Individually Guided Education of IGE is essentially a total system of education which takes into consideration the student\u27s previous knowledge, as well as how he goes about learning, how rapidly he learns, and other individual characteristics. Although the current thrust of Individually Guided Education-Secondary (IGE-S) is taking a different perspective than Individually Guided Education-Elemantary (IGE-E), the importance of establishing a positive learning climate or facilitative environment still appears to be an underlying concern. It would seem that such an environment would have the potential for affecting student self-esteem in a positive way. Because of these considerations, the basic research question in this study was stated as: 1. Will middle school students enrolled in a multi-unit Individually Guided Education program score significantly higher on a measure of self-esteem than middle school students enrolled in a traditional school? Secondary research questions which relate to self-esteem differences by reason of sex were also included in this study. They can be stated as: 2. Will middle schools boys score significantly higher on a measure of self-esteem than middle school girls regardless of program? 3. Will middle school boys enrolled in a multi-unit IGE program score significantly higher on a measure of self-esteem than middle school girls enrolled in the same type of program? 4. Will middle school boys enrolled in a traditional program score significantly higher on a measure of self-esteem than middle school girls enrolled in the same type of program?..

THE EFFECTIVENESS OF INDIVIDUALLY GUIDED EDUCATION ON THE DEVELOPMENT OF SELF-ESTEEM IN MIDDLE SCHOOL STUDENTS

Individually Guided Education of IGE is essentially a total system of education which takes into consideration the student\u27s previous knowledge, as well as how he goes about learning, how rapidly he learns, and other individual characteristics. Although the current thrust of Individually Guided Education-Secondary (IGE-S) is taking a different perspective than Individually Guided Education-Elemantary (IGE-E), the importance of establishing a positive learning climate or facilitative environment still appears to be an underlying concern. It would seem that such an environment would have the potential for affecting student self-esteem in a positive way. Because of these considerations, the basic research question in this study was stated as: 1. Will middle school students enrolled in a multi-unit Individually Guided Education program score significantly higher on a measure of self-esteem than middle school students enrolled in a traditional school? Secondary research questions which relate to self-esteem differences by reason of sex were also included in this study. They can be stated as: 2. Will middle schools boys score significantly higher on a measure of self-esteem than middle school girls regardless of program? 3. Will middle school boys enrolled in a multi-unit IGE program score significantly higher on a measure of self-esteem than middle school girls enrolled in the same type of program? 4. Will middle school boys enrolled in a traditional program score significantly higher on a measure of self-esteem than middle school girls enrolled in the same type of program?..

Providing Application-Level QoS In 3G/4G Wireless Systems: A Comprehensive Framework Based On Multi-Rate CDMA

The emerging applications for 3G and 4G wireless systems typically require highly heterogeneous and time-varying quality of service from the underlying protocol layers. The wireless links, however, provide only an unreliable communication channel that suffers from temporal outages. As a consequence, protocol mechanisms are needed, that based on the unreliable wireless links provide the different service qualities required by the emerging applications. In this paper we identify the emerging IP based applications for 3G and 4G wireless systems and categorize their QoS requirements. We discuss the wireless access mechanisms that show promise of being the basis for supporting these applications. We than propose a set of protocol mechanisms, that based on the discussed wireless access mechanisms, provide the required QoS for the different application categories