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Newport, WA, United States

Forrest L.A.,Ohio State University | Husein T.,Spokane Community College | Husein O.,Spokane Valley Ear
Laryngoscope | Year: 2012

Paradoxical vocal cord motion (PVCM), or vocal cord dysfunction, is a descriptive term for inappropriate adduction of the vocal folds during respiration. The laryngeal mistiming leads to breathing difficulty and is often misdiagnosed as refractory asthma. The etiology of PVCM has been unclear but has long been hypothesized to be psychological. The present thesis is a prospective study of 170 patients older than 18 years being evaluated for PVCM, with 117 of the 170 (68.8%) identified as having PVCM by video laryngoscopy. Laryngeal edema (P =.021) and reflux (P =.026) were increased in patients with PVCM. A flat inspiratory arm of the flow volume loop during spirometry testing was a predictor of PVCM (P =.034). A subgroup of 47 newly diagnosed patients with PVCM underwent psychological analysis. The psychological profiles were elucidated using the Minnesota Multiphasic Personality Inventory and the Life Experiences Survey to evaluate stress. Compared to established normative data, PVCM demonstrated a conversion disorder pattern (P <.01) but not an anxiety disorder or a correlation with stress. A subgroup, 11 of the 47 (23.4%), had normal psychological outcomes, and two of the 47 (4.3%) were identified as malingering. Previous studies have suggested that PVCM is strictly a psychological disorder. It is proposed that PVCM is a descriptive term that is multifactorial and the etiology should direct treatment. A classification scheme divides PVCM into primary, or psychological, and secondary. The secondary form consists of medical disorders divided into irritable larynx syndrome and neurologic disorders. © 2012 The American Laryngological, Rhinological, and Otological Society, Inc. Source

Kim K.-J.,Chung - Ang University | Lee Y.-A.,Chung - Ang University | Lee Y.-A.,National University of Singapore | Kim B.-J.,Chung - Ang University | And 7 more authors.
Cryobiology | Year: 2015

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. © 2015 Elsevier Inc. Source

Lee Y.-A.,Chung - Ang University | Lee Y.-A.,National University of Singapore | Kim Y.-H.,Chung - Ang University | Ha S.-J.,Chung - Ang University | And 8 more authors.
Journal of Animal Science | Year: 2014

Spermatogonial stem cells provide the foundation for continued adult spermatogenesis and their manipulation can facilitate assisted reproductive technologies or the development of transgenic animals. Because the pig is an important agricultural and biomedical research animal, the development of practical application techniques to manipulate the pig Spermatogonial stem cell is needed. The ability to preserve porcine Spermatogonial stem cell or testis tissue long term is one of these fundamental techniques. The objective of this study was to optimize methods to cryopreserve porcine Spermatogonial stem cell when freezing testis cells or testis tissue. To identify the most efficient cryopreservation technique, porcine testis cells (cell freezing) or testis tissue (tissue freezing) were frozen in medium containing dimethyl sulfoxide (DMSO) and fetal bovine serum (FBS) or DMSO, FBS, and various concentrations of trehalose (50, 100, or 200 mM). After thawing, undifferentiated germ cells were enriched and treatments were evaluated for cryopreservation efficiency. The tissue freezing method resulted in significantly greater germ cell recovery (P = 0.041) and proliferation capacity (P < 0.001) compared to the cell freezing treatment. Regardless of freezing method (cell vs. tissue), addition of 200 mM trehalose to freezing medium increased germ cell recovery and proliferation capacity compared to cells frozen using the same freezing method without trehalose. Interestingly, addition of trehalose to the tissue freezing medium significantly increased germ cell recovery (P = 0.012) and proliferation capacity (P = 0.004) compared to the cell freezing treatment supplemented with trehalose. To confirm that cryopreservation in trehalose improves the survival of Spermatogonial stem cell, testis cells enriched for undifferentiated germ cells were xenotransplanted into recipient mouse testes. Germ cells recovered from tissue frozen with 200 mM trehalose generated significantly more (P < 0.001) donor derived colonies than tissue frozen without trehalose. Regardless of cryopreservation medium or freezing method, testis cell recovery, viability, and proliferation capacity of germ cells after thawing were significantly lower compared to those of untreated fresh control. Nevertheless, these data demonstrate that undifferentiated porcine germ cells can be efficiently cryopreserved in the presence of 200 mM trehalose. © 2014 American Society of Animal Science. All rights reserved. Source

Kim K.-J.,Chung - Ang University | Cho C.M.,Chung - Ang University | Kim B.-G.,Chung - Ang University | Lee Y.-A.,Chung - Ang University | And 5 more authors.
Journal of Animal Science | Year: 2014

Undifferentiated germ cells have the capacity to develop into sperm capable of fertilizing oocytes and contributing genetic material to subsequent generations. The most primitive prepubertal undifferentiated germ cells include gonocytes and undifferentiated spermatogonia, including spermatogonial stem cells (SSC). Gonocytes, present in the testis at birth, differentiate into SSC, which maintain spermatogenesis for the remainder of the male's life. Because of their capacity to contribute to lifelong spermatogenesis, undifferentiated germ cells are attractive targets for genetic modification to produce transgenic animals, including cattle. To maximize the efficiency of genetic modification of bovine gonocytes and SSC, effective enrichment techniques need to be developed. Selection of bovine gonocytes using differential plating was improved 8-fold (P < 0.001) when using a combination of extracellular matrix proteins, including laminin, fibronectin, collagen type IV, and gelatin, compared to using laminin and gelatin alone. Selected cells labeled with PKH26 formed colonies of donor-derived germ cells after transplantation into recipient mouse testes, indicating putative stem cell function. Significantly more colonies (P < 0.001) per 1 × 105 viable transplanted cells were formed from isolated nonadherent cells (203 ± 23.2) compared to adherent (20 ± 2.7) or Percoll (45.5 ± 4.5) selected cells. After selection, some gonocytes were transduced using a lentiviral vector containing the transgene for the enhanced green fluorescent protein. Transduction efficiency was 17%. Collectively, these data demonstrate effective methods for the selection and genetic modification of bovine undifferentiated germ cells. © 2014 American Society of Animal Science. All rights reserved. Source

Kim Y.-H.,Chung - Ang University | Kim B.-J.,Chung - Ang University | Kim B.-G.,Chung - Ang University | Kim B.-G.,University of Pennsylvania | And 9 more authors.
Journal of Animal Science | Year: 2013

The objective of this study was to use fluorescence-activated cell sorting (FACS) and spermatogonial stem cell (SSC) xenotransplantation to identify cell surface markers of putative porcine SSC. Analysis of porcine testis cells enriched for spermatogonia using FACS indicated that nearly half of stage-specific embryonic antigen-1 (SSEA-1) expressing testis cells expressed the undifferentiated spermatogonia marker protein gene product 9.5 (PGP 9.5) whereas significantly fewer (P < 0.05) cells selected for thymus cell antigen-1 (Thy-1), also known as cluster of differentiation 90 (CD90), cluster of differentiation 9 (CD9), or other SSC markers expressed PGP 9.5. Immunocytochemical analysis indicated that promyelocytic leukemia zinc finger (PLZF) protein and germ cell lineage marker VASA homolog (VASA), also known as DEAD box protein 4 (DDX4), were expressed by SSEA-1 expressing germ cells. Spermatogonial stem cell xenotransplantation of testis cell populations enriched for cells expressing SSEA-1 generated significantly (P < 0.05; greater than 15-fold) more colonies of donor derived germ cells than unselected testis cells. In conclusion, these data indicate that SSC markers identified in rodents are likely not entirely conserved in pigs and that SSEA-1 is a marker for porcine undifferentiated spermatogonia including SSC in prepubertal boars and its expression may serve as a target for the further study of porcine germ cells. © 2013 American Society of Animal Science. All rights reserved. Source

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