Inhibitory effects of the nanoscale lysate derived from xenogenic dental pulp stem cells in lung cancer models.

BACKGROUND: Lung cancer is a highly prevalent malignancy and has the highest mortality rate among all tumors due to lymph node metastasis. Bone marrow and umbilical cord-derived mesenchymal stem cells (MSCs) have demonstrated tumor-suppressive effects on lung cancer. This study investigated the effects of DPSC lysate on proliferation, apoptosis, migration and invasion of cancer cells were studied in vivo and in vitro. METHODS: The proliferation, apoptosis, and migration/metastasis were evaluated by cell counting kit-8 assay, Annexin-V and propidium iodide staining, and the transwell assay, respectively. The expression levels of apoptosis-, cell cycle-, migration-, and adhesion-related mRNA and proteins were measured by qRT-PCR and western blot. The level and mRNA expression of tumor markers carcino embryonic antigen (CEA), neuron-specific enolase (NSE), and squamous cell carcinoma (SCC) were measured by Enzyme-linked immunosorbent assay (ELISA) and qRT-PCR. Finally, a tumor-bearing mouse model was constructed to observe the tumor-suppressive effect of DPSC lysate after intraperitoneal injection. RESULTS: DPSC lysate decreased the viability of A549 cells and induced apoptosis in lung cancer cells. Western blot confirmed that levels of Caspase-3, Bax, and Bad were increased, and Bcl-2 protein levels were decreased in A549 cells treated with DPSC lysate. In addition, DPSC lysate inhibited the migration and invasion of A549 cells; downregulated key genes of the cell cycle, migration, and adhesion; and significantly suppressed tumor markers. Xenograft results showed that DPSC lysate inhibited tumor growth and reduced tumor weight. CONCLUSIONS: DPSC lysate inhibited proliferation, invasion, and metastasis; promoted apoptosis in lung cancer cells; and suppressed tumor growth- potentially providing a cell-based alternative therapy for lung cancer treatment.

A new subtype of artificial cell-derived vesicles from dental pulp stem cells with the bioequivalence and higher acquisition efficiency compared to extracellular vesicles.

Extracellular vesicles (EVs) derived from dental pulp stem cells (DPSC) have been shown an excellent efficacy in a variety of disease models. However, current production methods fail to meet the needs of clinical treatment. In this study, we present an innovative approach to substantially enhance the production of 'Artificial Cell-Derived Vesicles (ACDVs)' by extracting and purifying the contents released by the DPSC lysate, namely intracellular vesicles. Comparative analysis was performed between ACDVs and those obtained through ultracentrifugation. The ACDVs extracted from the cell lysate meet the general standard of EVs and have similar protein secretion profile. The new ACDVs also significantly promoted wound healing, increased or decreased collagen regeneration, and reduced the production of inflammatory factors as the EVs. More importantly, the extraction efficiency is improved by 16 times compared with the EVs extracted using ultracentrifuge method. With its impressive attributes, this new subtype of ACDVs emerge as a prospective candidate for the future clinical applications in regenerative medicine.

Genome-scale CRISPR-Cas9 screening in stem cells: theories, applications and challenges.

Due to the rapid development of stem cell technology, there have been tremendous advances in molecular biological and pathological research, cell therapy as well as organoid technologies over the past decades. Advances in genome editing technology, particularly the discovery of clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-related protein 9 (Cas9), have further facilitated the rapid development of stem cell researches. The CRISPR-Cas9 technology now goes beyond creating single gene editing to enable the inhibition or activation of endogenous gene loci by fusing inhibitory (CRISPRi) or activating (CRISPRa) domains with deactivated Cas9 proteins (dCas9). These tools have been utilized in genome-scale CRISPRi/a screen to recognize hereditary modifiers that are synergistic or opposing to malady mutations in an orderly and fair manner, thereby identifying illness mechanisms and discovering novel restorative targets to accelerate medicinal discovery investigation. However, the application of this technique is still relatively rare in stem cell research. There are numerous specialized challenges in applying large-scale useful genomics approaches to differentiated stem cell populations. Here, we present the first comprehensive review on CRISPR-based functional genomics screening in the field of stem cells, as well as practical considerations implemented in a range of scenarios, and exploration of the insights of CRISPR-based screen into cell fates, disease mechanisms and cell treatments in stem cell models. This review will broadly benefit scientists, engineers and medical practitioners in the areas of stem cell research.