Proliferated Leydig Cells for Engineered Testis-like Tissue Regeneration with Testosterone-Secreting Ability

Hongda Bi,Xiaoyun Wang,Wei Liu,Yilin Cao,Guangdong Zhou,Xin Xing 한국조직공학과 재생의학회 2014 조직공학과 재생의학 Vol.11 No.5

Tissue engineering approach provides a hopeful strategy for reconstructing testis testosterone-secreting functions. However, limited source and low proliferative activity in vitro of Leydig cells (LCs, the main testosteroneproducing cells) makes testis-like tissue regeneration difficult to be achieved. This study explored the feasibility of in vitro expanding LCs and their potential application in testis-like tissue regeneration. LC lineage cells were isolated from Sprague-Dawley (SD) rats by differential adhesion method and cell composition was identified by expressions of 3β-HSD, LHR, LIFR, and c-kit. A modified expansion medium (EM) system was used to test the feasibility of in vitro expanding LC lineage. The results showed that the attached cells reached a high purification of LC lineage (>90%, indicated by positive expression of 3β-HSD) and that EM significantly enhanced proliferation of LC lineage compared to regular medium, which was testified to be related to the presence of stem LCs that was implied by positive expressions of LIFR and c-kit as well as the transition of 3β-HSD expression from negative to positive in partial cells. Importantly, the proliferated LCs showed relatively sustained testosterone-secreting ability in vitro and these cells combined with biodegradable scaffolds successfully regenerated testis-like tissue with sustained testosteronesecreting function in vivo, which was supported by the enhanced serum testosterone level in castrated rats. All these results indicated that the differential adhesion method could efficiently isolate and purify LC lineage and that EM system could efficiently promote proliferation and functional maintenance of LC lineage, providing a good cell source for testes-like tissue regeneration. Tissue engineering approach provides a hopeful strategy for reconstructing testis testosterone-secreting functions. However, limited source and low proliferative activity in vitro of Leydig cells (LCs, the main testosteroneproducing cells) makes testis-like tissue regeneration difficult to be achieved. This study explored the feasibility of in vitro expanding LCs and their potential application in testis-like tissue regeneration. LC lineage cells were isolated from Sprague-Dawley (SD) rats by differential adhesion method and cell composition was identified by expressions of 3β-HSD, LHR, LIFR, and c-kit. A modified expansion medium (EM) system was used to test the feasibility of in vitro expanding LC lineage. The results showed that the attached cells reached a high purification of LC lineage (>90%, indicated by positive expression of 3β-HSD) and that EM significantly enhanced proliferation of LC lineage compared to regular medium, which was testified to be related to the presence of stem LCs that was implied by positive expressions of LIFR and c-kit as well as the transition of 3β-HSD expression from negative to positive in partial cells. Importantly, the proliferated LCs showed relatively sustained testosterone-secreting ability in vitro and thesecells combined with biodegradable scaffolds successfully regenerated testis-like tissue with sustained testosteronesecreting function in vivo, which was supported by the enhanced serum testosterone level in castrated rats. All these results indicated that the differential adhesion method could efficiently isolate and purify LC lineage and that EM system could efficiently promote proliferation and functional maintenance of LC lineage, providing a good cell source for testes-like tissue regeneration.

Two Evaluation Budgets for the Measurement Uncertainty of Glucose in Clinical Chemistry

Hui Chen,Ling Zhang,Xiaoyun Bi,Xiaoling Deng 대한진단검사의학회 2011 Annals of Laboratory Medicine Vol.31 No.3

Background: Measurement uncertainty characterizes the dispersion of the quantity values attributed to a measurand. Although this concept was introduced to medical laboratories some years ago, not all medical researchers are familiar with it. Therefore, the evaluation and expression of measurement uncertainty must be highlighted using a practical example. Methods: In accordance with the procedure for evaluating and expressing uncertainty, provided by the Joint Committee for Guides in Metrology (JCGM), we used plasma glucose (Glu) as an example and defined it as the measurand. We then analyzed the main sources of uncertainty, evaluated each component of uncertainty, and calculated the combined uncertainty and expanded uncertainty with 2 budgets for single measurements and continuous monitoring, respectively. Results: During the measurement of Glu, the main sources of uncertainty included imprecision, within-subject biological variance (BVw), calibrator uncertainty, and systematic bias. We evaluated the uncertainty of each component to be 1.26%, 1.91%, 5.70%, 0.42%, and -2.87% for within-run imprecision, between-day imprecision, BVw, calibrator uncertainty, and systematic bias, respectively. For a single specimen, the expanded uncertainty was 7.38% or 6.1±0.45 mmol/L (κ=2); in continuous monitoring of Glu, the expanded uncertainty was 13.58% or 6.1±0.83 mmol/L (κ=2). Conclusions: We have demonstrated the overall procedure for evaluating and reporting uncertainty with 2 different budgets. The uncertainty is not only related to the medical laboratory in which the measurement is undertaken, but is also associated with the calibrator uncertainty and the biological variation of the subject. Therefore, it is helpful in explaining the accuracy of test results. Background: Measurement uncertainty characterizes the dispersion of the quantity values attributed to a measurand. Although this concept was introduced to medical laboratories some years ago, not all medical researchers are familiar with it. Therefore, the evaluation and expression of measurement uncertainty must be highlighted using a practical example. Methods: In accordance with the procedure for evaluating and expressing uncertainty, provided by the Joint Committee for Guides in Metrology (JCGM), we used plasma glucose (Glu) as an example and defined it as the measurand. We then analyzed the main sources of uncertainty, evaluated each component of uncertainty, and calculated the combined uncertainty and expanded uncertainty with 2 budgets for single measurements and continuous monitoring, respectively. Results: During the measurement of Glu, the main sources of uncertainty included imprecision, within-subject biological variance (BVw), calibrator uncertainty, and systematic bias. We evaluated the uncertainty of each component to be 1.26%, 1.91%, 5.70%, 0.42%, and -2.87% for within-run imprecision, between-day imprecision, BVw, calibrator uncertainty, and systematic bias, respectively. For a single specimen, the expanded uncertainty was 7.38% or 6.1±0.45 mmol/L (κ=2); in continuous monitoring of Glu, the expanded uncertainty was 13.58% or 6.1±0.83 mmol/L (κ=2). Conclusions: We have demonstrated the overall procedure for evaluating and reporting uncertainty with 2 different budgets. The uncertainty is not only related to the medical laboratory in which the measurement is undertaken, but is also associated with the calibrator uncertainty and the biological variation of the subject. Therefore, it is helpful in explaining the accuracy of test results.