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This paper presents a new integrated microfluidic chip that automatically performs ribonucleic acid (RNA) extraction and reverse transcription (RT) processes. The microfluidic system consists of a microfluidic control module and a magnetic bio-separator. The microfluidic control module can perform pumping and mixing of small amount of fluids and subsequent purification and concentration of RNA samples by incorporating with the magnetic bio-separator consisting of 2-dimension twisted microcoils. Notably, the magnetic bio-separators are developed either to generate the required magnetic field to perform the separation of magnetic beads or to work as a micro-heater to control the temperature field for the following RT process. Experimental results show that the total RNA can be successfully purified and extracted by using magnetic beads and the subsequent RT processing of the RNA can be performed automatically. Total RNA is successfully extracted and purified from T98 cells utilizing the microfluidic system, which is comparable with the conventional methods. The whole automatic procedure of RNA sample extraction only takes 35 min, which is much faster than the conventional method (more than 2 h). As a whole, the developed microfluidic system may provide a powerful platform for rapid RNA extraction and RT processes for further biomedical applications.

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Chronic arsenic exposure has been linked to endothelial dysfunction and apoptosis. We investigate the involvement of unfolded protein response (UPR) signaling in the arsenic-mediated cytotoxicity of the SVEC4-10 mouse endothelial cells. The SVEC4-10 cells underwent apoptosis in response to As2O3 dose- and time-dependently, accompanied by increased accumulation of calcium, and activation of caspase-3. These phenomena were completely inhibited by α-lipoic acid (LA), which did not scavenge ROS over-production, but were only partially or not ameliorated by tiron, a potent superoxide scavenger. Moreover, arsenic activated UPR, leading to phosphorylation of eukaryotic translation initiation factor 2 subunit α (eIF2α), induction of ATF4, and processing of ATF6. Treatment with arsenic also triggered the expression of endoplasmic reticulum (ER) stress markers, GRP78 (glucose-regulated protein), and CHOP (C/EBP homologous protein). The activation of eIF2α, ATF4 and ATF6 and expression of GRP78 and CHOP are repressed by both LA and tiron, indicating arsenic-induced UPR is mediated through ROS-dependent and ROS-independent pathways. Arsenic also induced ER stress-inducible genes, BAX, PUMA (p53 upregulated modulator of apoptosis), TRB3 (tribbles-related protein 3), and SNAT2 (sodium-dependent neutral amino acid transporter 2). Consistent with intracellular calcium and cell viability data, ROS may not be important in arsenic-induced death, because tiron did not affect the expression of these pro-apoptotic genes. In addition, pretreatment with salubrinal, a selective inhibitor of eIF2α dephosphorylation, enhanced arsenic-induced GRP78 and CHOP expression and partially prevented arsenic cytotoxicity in SVEC4-10 cells. Taken together, these results suggest that arsenic-induced endothelial cytotoxicity is associated with ER stress, which is mediated by ROS-dependent and ROS-independent signaling.

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