Cryopreservation of putative pre-pubertal bovine spermatogonial stem cells by slow freezing.

Development of techniques for the preservation of mammalian spermatogonial stem cells (SSCs) is a critical step in commercial application of SSC based technologies, including species preservation, amplification of agriculturally valuable germ lines, and human fertility preservations. The objective of this study was to develop an efficient cryopreservation protocol for preservation of bovine SSCs using a slow freezing technique. To maximize the efficiency of SSC cryopreservation, the effects of various methods (tissue vs. cell freezing) and cryoprotective agents (trehalose, sucrose, and polyethylene glycol [PEG]) were tested. Following thawing, cells were enriched for undifferentiated spermatogonia by differential plating and evaluated for recovery rate, proliferation capacity, and apoptosis. Additionally, putative stem cell activity was assessed using SSC xenotransplantation. The recovery rate, and proliferation capacity of undifferentiated spermatogonia were significantly greater for germ cells frozen using tissue freezing methods compared to cell freezing methods. Cryopreservation in the presence of 200 mM trehalose resulted in significantly greater recovery rate, proliferation capacity, and apoptosis of germ cells compared to control. Furthermore, cryopreservation using the tissue freezing method in the presence of 200 mM trehalose resulted in the production of colonies of donor-derived germ cells after xenotransplantation into recipient mouse testes, indicating putative stem cell function. Collectively, these data indicate that cryopreservation using tissue freezing methods in the presence of 200 mM trehalose is an efficient cryopreservation protocol for bovine SSCs.

Modified folding radial forearm flap in soft palate and tonsillar fossa reconstruction.

Wide excision of cancer arising from the tonsillar fossa and soft palate has several functional sequelae (e.g., speech, swallowing, chewing, and breathing) that require surgical restoration of the pharyngeo-palatal structure and optimal velopharyngeal function. For this purpose, several kinds of surgical procedures have been introduced. Our method to reconstruct the tonsillar fossa and soft palate entails folding the flaps and reconstructions at the same time as the oral and nasal planes, with some modifications.Patient 1 was a 64-year-old man with left soft palate cancer. After wide excision of the tumor, the defect size of the nasal floor was 3 × 3 cm, and that of soft palate and tonsillar fossa was 8 × 5 cm. Patient 2 was a 49-year-old man with left tonsil cancer. The defect size of the nasal floor was 3 × 3 cm, and that of left lateral wall of the tonsillar fossa was 8 × 3 cm. For reconstruction of oral, nasal, and tonsillar plane, we designed the flap fit to the defect site, especially cutting of the edge of the square plane of the flap to a round shape.Both patients achieved good functional recovery without surgical complications. The average speech intelligibility score in the 2 patients was 10. Swallowing functional score was 4 in both patients. Creative reconstruction with modified radial forearm free flap for tonsillar and soft palate area makes it possible to restore velopharyngeal function to levels close to the preoperative condition.

Therapeutic effects of a liquid bandage prepared with cellulose powders from Styela clava tunics and Broussonetia kazinoki bark: Healing of surgical wounds on the skin of Sprague Dawley rats.

Cellulose in different forms has extensively been applied in biomedical treatments, including scaffolding, tissue engineering and tissue formation. To evaluate the therapeutic effects of a liquid bandage (LB) prepared with cellulose powders from Styela clava tunics (SCT) and Broussonetia kazinoki bark (BSLB) for healing cutaneous wounds, the remedial effects of a low concentration (LoBSLB) and a high concentration (HiBSLB) of BSLB on skin regeneration and toxicity in Sprague Dawley rats. Results indicated that the total area of skin involved in the surgical wound was lower in the BSLB­treated group compared with the Vehicle­treated group at days 4­12, although some variations were observed in the HiBSLB­treated group. In addition, the BSLB­treated group showed significantly enhanced width of the re­epithelialization region and epidermal thickness when compared with the Vehicle­treated group. Furthermore, significant stimulation in the expression level of collagen­1 and the signaling pathway of VEGF after topical application of BSLB was indicated. No liver or kidney toxicities were detected for either doses of BSLB. Overall, the results of the present study suggest that BSLB accelerates the process of wound healing in surgical skin wounds of Sprague Dawley rats through stimulation of re­epithelialization and connective tissue formation, without any accompanying significant toxicity.

Hypercapnia does not shorten emergence time from propofol anesthesia: a pilot randomized clinical study.

BACKGROUND: The elimination of anesthetic agents is a decisive factor in the emergence from general anesthesia. In this pilot study, we hypothesized that hypercapnia would decrease the emergence time from propofol anesthesia by increasing cardiac output and cerebral blood flow. METHODS: A total of 32 patients were randomly divided into two groups based on the end-tidal carbon dioxide values: 30 mmHg (the hypocapnia group) and 50 mmHg (the hypercapnia group). Propofol and remifentanil were infused to maintain a bispectral index of 40-50. Remifentanil infusion was stopped 10 min before the discontinuation of propofol. After cessation of propofol infusion, ventilation settings in the hypocapnia group were maintained constant; a rebreathing tube was connected to the respiratory circuit in the hypercapnia group. The time to spontaneous respiration, eye opening (primary endpoint), mouth opening, and tracheal extubation was recorded and analyzed. RESULTS: Time to eye opening was 9.7 (1.3) min in the hypocapnia group and 9.0 (1.0) min in the hypercapnia group. The difference in the mean times to eye opening between groups was -0.7 min (95% CI, -4.0 to 2.7, P = 0.688). On multiple regression analysis, there was a significant difference in the mean time to eye opening between males and females. Females recovered about 3.6 min faster than males (95% CI, -6.1 to -1.1, P = 0.009). CONCLUSIONS: We could not detect a beneficial effect of hypercapnia on propofol emergence time. Irrespective of hypercapnia, females seemed to recover faster than males.

Detailed anatomy of the retaining ligaments of the mandible for facial rejuvenation.

PURPOSE: To present anatomical characteristics of three true ligaments of the mandible and better understanding of jowl formation. MATERIALS AND METHODS: Facial dissections were performed in 10 fresh cadavers (20 hemifaces) using 2.5 magnifying surgical loupes. Mental ligament, medial mandibular ligament, and mandibular osteocutaneous ligament were identified as thick fibrous tissue originating from the mandible. RESULTS: The location of the mental ligament was 9.1 ± 2.8 mm lateral to the midline and 8.4 ± 3.8 mm superior to the inferior border of the mandible. The medial mandibular ligament was located 28.5 ± 4.9 mm lateral to the midline and 8.4 ± 2.7 mm superior to the inferior border of the mandible. The mandibular osteocutaneous ligament was located 51.1 ± 4.5 mm lateral to the midline and 9.1 ± 3.1 mm superior to the inferior border of the mandible. CONCLUSIONS: We investigated location and tension of two unfamiliar ligaments in the mandible, the mental and medial mandibular ligaments medial to the mandibular osteocutaneous ligament, and confirmed them as true ligaments by histology. Our findings suggest that the cause of the jowl formation is mainly descent of the cheek fat compartments, but not laxity of mandibular osteocutaneous ligament.