TY - JOUR
T1 - Laser-assisted cryosurgery in ex vivo mice hepatic tissue
T2 - Viability assays using green fluorescent protein
AU - Martínez-Suástegui, L.
AU - Duperray, B.
AU - Godinez, F.
AU - Guillén, G.
AU - Slade, A.
AU - Aguilar, G.
N1 - Funding Information:
G. Aguilar and L. Martínez-Suástegui would like to acknowledge the support of the UC MEXUS-CONACYT program for a Postdoctoral Fellowship granted to the latter to spend a year as a postdoctoral researcher at UCR. The authors thank David R. Hennings CoolTouch CEO for providing the laser system and optical fibers. B. Duperray acknowledges the school of ENSICaen and F. Godinez acknowledges the UCR Undergraduate Research Grant, the UC-LEADS, and the CAMP programs for the support provided to him. G. Guillén acknowledges support of the DGAPA program (UNAM) for a Fellowship to spend an internship at UCR. The authors thank Feng Sun and Florian Langer for their help and efforts with H&E staining. In particular, the authors appreciate the input of A.M. Walker from the University of California Riverside, Division of Biomedical Sciences and S. Robert from the VIB Department of Plant Systems Biology, Ghent University. The authors gratefully acknowledge expert animal husbandry by Ms. Sally Scott. This work was supported in part by NSF under grant NSF-CTS-SGER: 0609662.
PY - 2011/2
Y1 - 2011/2
N2 - An experimental investigation is carried out to develop a novel approach to cryosurgery, where laser heating counteracts tissue freezing to better confine damage to the targeted cancerous tissue within a lethal low-temperature isothermal boundary-an approach we refer to as laser-assisted cryosurgery (LAC). The advantage of this procedure relative to conventional cryosurgery assisted with urethral warmers or cryoheaters is that laser heating provides volumetric rather than superficial heating, which leads to deeper penetration, more homogeneous tissue protection and better demarcation of the destructive freezing effect to a well-defined targeted volume. Tissue viability assays are performed using green fluorescence protein (GFP) as a viability marker and correlated with temperature history after performing LAC procedures on ex vivo mice hepatic tissue. The limit for cell denaturation at the irradiated surface predicted by GFP analysis is further confirmed using reverse transcription polymerase chain reaction (RT-PCR). In addition, the correlation between GFP fluorescence and cell viability and loss of GFP fluorescence in non-viable cells has been tested and validated by histological analysis using a standard cell viability measuring method (hematoxylin and eosin staining). Analysis of our experimental measurements show that reproducible thermal gradients (of 236 °C/cm) and predictable tissue necrosis can be reliably produced by LAC without exceeding temperature thresholds for cell denaturation (of T surf ≈ 48 °C) beyond preset tissue boundaries (with resolution of 0.1 °C/mm). The results have shown the feasibility of controlling temperatures at specified tissue locations to prevent hyperthermal or freezing damage.
AB - An experimental investigation is carried out to develop a novel approach to cryosurgery, where laser heating counteracts tissue freezing to better confine damage to the targeted cancerous tissue within a lethal low-temperature isothermal boundary-an approach we refer to as laser-assisted cryosurgery (LAC). The advantage of this procedure relative to conventional cryosurgery assisted with urethral warmers or cryoheaters is that laser heating provides volumetric rather than superficial heating, which leads to deeper penetration, more homogeneous tissue protection and better demarcation of the destructive freezing effect to a well-defined targeted volume. Tissue viability assays are performed using green fluorescence protein (GFP) as a viability marker and correlated with temperature history after performing LAC procedures on ex vivo mice hepatic tissue. The limit for cell denaturation at the irradiated surface predicted by GFP analysis is further confirmed using reverse transcription polymerase chain reaction (RT-PCR). In addition, the correlation between GFP fluorescence and cell viability and loss of GFP fluorescence in non-viable cells has been tested and validated by histological analysis using a standard cell viability measuring method (hematoxylin and eosin staining). Analysis of our experimental measurements show that reproducible thermal gradients (of 236 °C/cm) and predictable tissue necrosis can be reliably produced by LAC without exceeding temperature thresholds for cell denaturation (of T surf ≈ 48 °C) beyond preset tissue boundaries (with resolution of 0.1 °C/mm). The results have shown the feasibility of controlling temperatures at specified tissue locations to prevent hyperthermal or freezing damage.
KW - Cryoablation
KW - Prostate cancer
KW - RT-PCR
UR - http://www.scopus.com/inward/record.url?scp=79951557287&partnerID=8YFLogxK
U2 - 10.1007/s10439-010-0186-0
DO - 10.1007/s10439-010-0186-0
M3 - Artículo
C2 - 20963494
SN - 0090-6964
VL - 39
SP - 636
EP - 648
JO - Annals of Biomedical Engineering
JF - Annals of Biomedical Engineering
IS - 2
ER -