In-vivo optical imaging of hsp70 expression to assess collateral tissue damage associated with infrared laser ablation of skin

Abstract

Laser surgical ablation is achieved by selecting laser parameters that remove confined volumes of target tissue and cause minimal collateral damage. Previous studies have measured the effects of wavelength on ablation, but neglected to measure the cellular impact of ablation on cells outside the lethal zone. In this study, we use optical imaging in addition to conventional assessment techniques to evaluate lethal and sublethal collateral damage after ablative surgery with a free-electron laser (FEL). Heat shock protein (HSP) expression is used as a sensitive quantitative marker of sublethal damage in a transgenic mouse strain, with the hsp70 promoter driving luciferase and green fluorescent protein (GFP) expression (hsp70A1-L2G). To examine the wavelength dependence in the mid-IR, laser surgery is conducted on the hsp70A1-L2G mouse using wavelengths targeting water (OH stretch mode, 2.94 μm), protein (amide-II band, 6.45 μm), and both water and protein (amide-I band, 6.10 μm). For all wavelengths tested, the magnitude of hsp70 expression is dose-dependent and maximal 5 to 12 h after surgery. Tissues treated at 6.45 μm have approximately 4× higher hsp70 expression than 6.10 μm. Histology shows that under comparable fluences, tissue injury at the 2.94-μm wavelength was 2× and 3× deeper than 6.45 and 6.10 μm, respectively. The 6.10-μm wavelength generates the least amount of epidermal hyperplasia. Taken together, this data suggests that the 6.10-μm wavelength is a superior wavelength for laser ablation of skin.

Invited Review Article: Current State of Research on Biological Effects of Terahertz Radiation

“…For example, cells exposed to 45°C for 15 min were observed to undergo transient G 2 block, while when exposed to 45°C for 30 min, they exhibited a long lasting G 2 phase and a late S-phase block [73]. The CSR mechanism is well documented in the literature [66][67][68][69][74][75][76][77], and the general consensus is that cells activate these mechanisms to combat and survive hyperthermic stress. CSR mechanisms involve many intracellular pathways, including redox, DNA sensing and repair, molecular chaperones, proteolysis, energy metabolism, and apoptosis [78].…”

“…These proteins, collectively referred to as minimal stress proteins, are significantly upregulated by cells immediately after exposure to stress. The most widely studied family of minimal stress proteins are the heat shock proteins (Hsps), which includes Hsp70, Hsp40, Hsp60, and Hsp105 [66][67][68][69][74][75][76][77] (Fig. 4a, c).…”

“…The most common thermal effects observed at a cellular level include the following: (i) stimulation of cell growth and metabolism; (ii) morphological changes (i.e., swelling, blebbing, shrinking); (iii) activation of cellular stress response (CSR) mechanisms [66][67][68][69], and (iv) cell death via apoptotic and necrotic pathways [66][67][68][69]. The thermal sensitivity of cells is cell type dependent; however, all cell lines typically share a similar response trend.…”

Pulse duration determines levels of Hsp70 induction in tissues following laser irradiation

“…which supports the idea of a graded response corresponding to the extent of stress in this region. This mouse line has also been used to reveal Hsp70 expression patterns following the laser thermal stress/damage using a free electron laser at 2.94, 6.1, and 6.45 μm in wavelength 22 and a 1.85 μm pulsed diode laser. 23 In addition to laser related research, this mouse line has also been used to study the expression of Hsp70 following a high intensity focused ultrasound stimulation in vivo.…”

Development of a laser-activated mesoporous silica nanocarrier delivery system for applications in molecular and genetic research

“…33,42 Previous reports have illustrated spatially controlled laser-activated HSP promoter-induced genes in transgenic models. [43][44][45] However, to the extent of our knowledge, the use of an HSP construct as cargo for MSNPs that can be thermally triggered by IR laser irradiation to increase expression has not yet been reported. We thus investigated, for the first time, the use of IR laser irradiation of cells containing HSP cargo-loaded MSNPs as a method of inducing a cellular heat-shock response to specifically increase expression of the cargo.…”