In vitro fertilization and subsequent development of porcine oocytes using cryopreserved and liquid-stored spermatozoa from various boars

We had previously developed a porcine IVF system using a chemically defined medium, i.e., porcine gamete medium supplemented with theophylline, adenosine, and cysteine (PGMtac). In the present study, we investigated the utility of this IVF system using different types of semen: (1) cryopreserved ejaculated (n = 8); (2) cryopreserved epididymal (n = 4); and (3) liquid-stored ejaculated (n = 5). Cryopreserved spermatozoa were prepared by three methods. In vitro-matured porcine oocytes were fertilized for 20 h in PGMtac using each type of semen, and the presumptive zygotes were cultured in porcine zygote medium (PZM)-4 for 5 days. In the case of frozen-thawed spermatozoa, the number of spermatozoa per penetrated oocyte (1.1-1.7), rate of blastocyst formation (26-56%), and total number of cells per blastocyst (34-49) differed (P < 0.05) among freezing methods. However, blastocysts were produced using all types of cryopreserved spermatozoa (14-75%). When spermatozoa were liquid-stored for 1-14 days after semen collection, the rate of sperm penetration (P < 0.05) decreased as storage time increased, although there was no significant reduction in sperm motility during storage. In all groups, semen that had been stored within 10 days after collection enabled blastocyst production in vitro (20-48%). In conclusion, this IVF system, which uses a chemically defined medium, had widespread utility with both frozen-thawed and liquid-stored spermatozoa.     

Change in content of sugars and free amino acids in potato tubers under short-term storage at low temperature and the effect on acrylamide level after frying

Changes in the sugar and amino acid contents of potato tubers during short-term storage and the effect on the acrylamide level in chips after frying were investigated. The acrylamide content in chips began to increase after 3 days of storage at 2 degrees C in response to the increase of glucose and fructose contents in the tubers. There was strong correlation between the reducing sugar content and acrylamide level, R(2)=0.873 for fructose and R(2)=0.836 for glucose. The sucrose content had less correlation with the acrylamide content because of its decrease after 4 weeks of storage at 2 degrees C, while the reducing sugar in potato tubers and the acrylamide in chips continued to increase. The contents of the four amino acids, i.e., asparatic acid, asparagine, glutamic acid and glutamine, showed no significant correlation with the acrylamide level. These results suggest that the content of reducing sugars in potato tubers determined the degree of acrylamide formation in chips. The chip color, as evaluated by L* (lightness), was correlated well with the acrylamide content.     

Facilitated production of secretory IgA against Shiga toxin B subunits by intranasal application of antigen-coated polystyrene microspheres

We examined the effects of microspheres as antigen carriers in mucosal immunization. Shiga toxin B subunits (Stx1B) were adsorbed on 6 mum polystyrene microspheres, which were then intranasally administered to mice together with cholera toxin (CT). Stx1B-specific serum IgG production and secretory IgA production at local mucosal sites were enhanced by the use of microspheres. When OVA was used as a model antigen, secretory IgA production but not serum IgG production was enhanced on the use of microspheres. These results indicated that microspheres provide a useful means of potentiating the immune response against Stx1B with weak immunogenicity.     

A serum factor capable of stimulating hyaluronic acid synthesis in cultured rat fibroblasts.

Calf serum as well as rat and mouse sera has a factor that stimulates hyaluronic acid synthesis in cultured rat fibroblasts. Such a factor was partially purified from calf serum and characterized. It has a molecular weight of approximately 150,000. The activity of the factor is lost by treatment with pronase and by periodate oxidation. It is suggested, therefore, that the factor is a glycoprotein. Its susceptibility to alpha-mannosidase and affinity for Con A-Sepharose may suggest that the factor contains a mannose residue(s) which is essential for the activity to induce hyaluronic acid synthesis.     

Impact of Lymph Node Ratio on Oncologic Outcomes in ypStage III Rectal Cancer Patients Treated with Neoadjuvant Chemoradiotherapy followed by Total Mesorectal Excision,and Postoperative Adjuvant Chemotherapy

Purpose

To evaluate the prognostic impact of the lymph node ratio (LNR) in ypStage III rectal cancer patients who were treated with neoadjuvant chemoradiotherapy (NCRT).

Materials and Methods

We retrospectively reviewed the data of 638 consecutive patients who underwent NCRT followed by total mesorectal excision, and postoperative adjuvant chemotherapy for rectal cancer from 2004 to 2011. Of these, 125 patients were positive for lymph node (LN) metastasis and were analyzed in this study.

Results

The median numbers of examined and metastatic LNs were 17 and 2, respectively, and the median LNR was 0.143 (range, 0.02–1). Median follow-up time was 55 months. In multivariate analyses, LNR was an independent prognostic factor for overall survival (OS) (hazard ratio [HR] 2.17, p = 0.041), disease-free survival (DFS) (HR 2.28, p = 0.005), and distant metastasis-free survival (DMFS) (HR 2.30, p = 0.010). When ypN1 patients were divided into low (low LNR ypN1 group) and high LNR (high LNR ypN1 group) according to a cut-off value of 0.152, the high LNR ypN1 group had poorer OS (p = 0.043) and DFS (p = 0.056) compared with the low LNR ypN1 group. And there were no differences between the high LNR ypN1 group and the ypN2 group in terms of the OS (p = 0.703) and DFS (p = 0.831).

Conclusions

For ypN-positive rectal cancer patients, the LNR was a more effective prognostic marker than the ypN stage, circumferential resection margin, or tumor regression grade after NCRT, and could be used to discern the high-risk group among ypN1 patients.     

Sex Differences in Object Manipulation in Wild Immature Chimpanzees (Pan troglodytes schweinfurthii) and Bonobos (Pan paniscus): Preparation for Tool Use?

Sex differences in immatures predict behavioural differences in adulthood in many mammal species. Because most studies have focused on sex differences in social interactions, little is known about possible sex differences in ‘preparation’ for adult life with regards to tool use skills. We investigated sex and age differences in object manipulation in immature apes. Chimpanzees use a variety of tools across numerous contexts, whereas bonobos use few tools and none in foraging. In both species, a female bias in adult tool use has been reported. We studied object manipulation in immature chimpanzees at Kalinzu (Uganda) and bonobos at Wamba (Democratic Republic of Congo). We tested predictions of the ‘preparation for tool use’ hypothesis. We confirmed that chimpanzees showed higher rates and more diverse types of object manipulation than bonobos. Against expectation, male chimpanzees showed higher object manipulation rates than females, whereas in bonobos no sex difference was found. However, object manipulation by male chimpanzees was play-dominated, whereas manipulation types of female chimpanzees were more diverse (e.g., bite, break, carry). Manipulation by young immatures of both species was similarly dominated by play, but only in chimpanzees did it become more diverse with age. Moreover, in chimpanzees, object types became more tool-like (i.e., sticks) with age, further suggesting preparation for tool use in adulthood. The male bias in object manipulation in immature chimpanzees, along with the late onset of tool-like object manipulation, indicates that not all (early) object manipulation (i.e., object play) in immatures prepares for subsistence tool use. Instead, given the similarity with gender differences in human children, object play may also function in motor skill practice for male-specific behaviours (e.g., dominance displays). In conclusion, even though immature behaviours almost certainly reflect preparation for adult roles, more detailed future work is needed to disentangle possible functions of object manipulation during development.     

Orbit/CLASP Is Required for Myosin Accumulation at the Cleavage Furrow in Drosophila Male Meiosis

Peripheral microtubules (MTs) near the cell cortex are essential for the positioning and continuous constriction of the contractile ring (CR) in cytokinesis. Time-lapse observations of Drosophila male meiosis showed that myosin II was first recruited along the cell cortex independent of MTs. Then, shortly after peripheral MTs made contact with the equatorial cortex, myosin II was concentrated there in a narrow band. After MT contact, anillin and F-actin abruptly appeared on the equatorial cortex, simultaneously with myosin accumulation. We found that the accumulation of myosin did not require centralspindlin, but was instead dependent on Orbit, a Drosophila ortholog of the MT plus-end tracking protein CLASP. This protein is required for stabilization of central spindle MTs, which are essential for cytokinesis. Orbit was also localized in a mid-zone of peripheral MTs, and was concentrated in a ring at the equatorial cortex during late anaphase. Fluorescence resonance energy transfer experiments indicated that Orbit is closely associated with F-actin in the CR. We also showed that the myosin heavy chain was in close proximity with Orbit in the cleavage furrow region. Centralspindlin was dispensable in Orbit ring formation. Instead, the Polo-KLP3A/Feo complex was required for the Orbit accumulation independently of the Orbit MT-binding domain. However, orbit mutations of consensus sites for the phosphorylation of Cdk1 or Polo did not influence the Orbit accumulation, suggesting an indirect regulatory role of these protein kinases in Orbit localization. Orbit was also necessary for the maintenance of the CR. Our data suggest that Orbit plays an essential role as a connector between MTs and the CR in Drosophila male meiosis.