Facilitation of axonal transcriptome analysis with quantitative microfluidic devices.

Microfluidic chambers are powerful tools for studying axonal mRNA localization and translation in neurons. In addition to specific manipulation and measurements of axons, microfluidic chambers are used for collecting axonal materials to perform axonal transcriptome analysis. However, traditional bipartite and tripartite chambers have limitations either in purity or quantity of collected axons. Here, we improved the design of traditional chambers. Moreover, we developed two new quantitative chambers, multi-compartmental quantitative bipartite chamber (MQBC) and long quantitative tripartite chamber (LQTC). Compared with the traditional chambers, MQBC and LQTC could dramatically increase the efficiency in collecting axonal RNA. Finally, we applied these chambers to do comparative axon transcriptome analysis of different types of neurons. Thus, our newly designed quantitative chambers significantly improve axon collection efficiency and facilitate axonal transcriptome analysis.

The m6 A Readers YTHDF1 and YTHDF2 Synergistically Control Cerebellar Parallel Fiber Growth by Regulating Local Translation of the Key Wnt5a Signaling Components in Axons.

Messenger RNA m6 A modification is shown to regulate local translation in axons. However, how the m6 A codes in axonal mRNAs are read and decoded by the m6 A reader proteins is still unknown. Here, it is found that the m6 A readers YTHDF1 and YTHDF2 are both expressed in cerebellar granule cells (GCs) and their axons. Knockdown (KD) of YTHDF1 or YTHDF2 significantly increases GC axon growth rates in vitro. By integrating anti-YTHDF1&2 RIP-Seq with the quantitative proteomic analysis or RNA-seq after KD of YTHDF1 or YTHDF2, a group of transcripts which may mediate the regulation of GC axon growth by YTHDFs is identified. Among them, Dvl1 and Wnt5a, encoding the key components of Wnt pathway, are further found to be locally translated in axons, which are controlled by YTHDF1 and YTHDF2, respectively. Specific ablation of Ythdf1 or Ythdf2 in GCs increases parallel fiber growth, promotes synapse formation in cerebellum in vivo, and improves motor coordination ability. Together, this study identifies a mechanism by which the m6 A readers YTHDF1 and YTHDF2 work synergistically on the Wnt5a pathway through regulating local translation in GC axons to control cerebellar parallel fiber development.

The Application of Microfluidic Technologies in Aptamer Selection.

Aptamers are sequences of single-strand oligonucleotides (DNA or RNA) with potential binding capability to specific target molecules, which are increasingly used as agents for analysis, diagnosis, and medical treatment. Aptamers are generated by a selection method named systematic evolution of ligands by exponential enrichment (SELEX). Numerous SELEX methods have been developed for aptamer selections. However, the conventional SELEX methods still suffer from high labor intensity, low operation efficiency, and low success rate. Thus, the applications of aptamer with desired properties are limited. With their advantages of low cost, high speed, and upgraded extent of automation, microfluidic technologies have become promising tools for rapid and high throughput aptamer selection. This paper reviews current progresses of such microfluidic systems for aptamer selection. Comparisons of selection performances with discussions on principles, structure, operations, as well as advantages and limitations of various microfluidic-based aptamer selection methods are provided.

The Pharmacological Effects of Spatholobi Caulis Tannin in Cervical Cancer and Its Precise Therapeutic Effect on Related circRNA.

The chemical components of Spatholobi Caulis tannin (SCT) have a modest therapeutic effect in patients with cervical cancer. However, the active components and the mechanism of action of SCT in HeLa cervical cancer cells need to be further studied. In this paper, 3D microfluidic chip technology was applied to simulate the effects of tannins in the human body, and the appropriate dose and time of administration were calculated. The cell cycle and apoptosis experiments demonstrated that SCT inhibits proliferation and stimulated apoptosis in HeLa cells. The differentially expressed genes were screened using The Cancer Genome Atlas (TCGA) and the GEO databases to identify common differentially expressed genes. A bioinformatic analysis of relevant genes, analysis using the molecular docking technique, and survival analysis were used to predict the target genes of SCT. Circular RNAs (circRNAs) associated with the SCT target genes and the regulatory effects of SCT on these circRNAs were determined. These studies showed that SCT mediates related circRNAs in HeLa cells to inhibit proliferation and promote apoptosis in HeLa cells. Thus, SCT may be an effective strategy for treating cervical cancer.

3D microfluidic in vitro model and bioinformatics integration to study the effects of Spatholobi Caulis tannin in cervical cancer.

Cervical cancer is considered the fourth most common malignant disease in women. Recently, tannin from Spatholobi Caulis (TTS) has been shown to have potent anticancer and antiproliferative characteristics in a few preliminary studies. This experiment used 3D microfluidic, flow cytometry, and gene chip technology to study the efficacy and mechanism of action of TTS, as well as molecular docking technology to study the effect of drugs on related proteins. The cell survival rates of the five groups measured by the 3D microfluidic chip were 94%, 85%, 64%, 55%, and 42%, respectively. With the increase in drug concentration, the cell survival rate gradually decreased. Apoptosis rates detected in the five groups were 2.12%, 15.87%, 33.40%, 41.13%, and 55.10%, respectively. These data suggest that TTS can promote cell apoptosis. The percentages of cells in the G0/G1 phase were 43.39%, 55.07%, 59.57%, 64.56%, and 67.39% in the five groups, respectively. TTS was demonstrated to inhibit the conversion of cells from G0/G1 to S phase and G2/M phase and inhibit gene and protein synthesis to block cell proliferation. TTS can effectively modulate pathogenic proteins. The results confirmed the efficacy of TTS against HeLa cells and that TTS can be used as an adjunct in cervical cancer prevention and treatment.