Exosomes are attractive as nucleic-acid carriers because of their favourable pharmacokinetic and immunological properties and their ability to penetrate physiological barriers that are impermeable to synthetic drug-delivery vehicles. However, inserting exogenous nucleic acids, especially large messenger RNAs, into cell-secreted exosomes leads to low yields. Here we report a cellular-nanoporation method for the production of large quantities of exosomes containing therapeutic mRNAs and targeting peptides. We transfected various source cells with plasmid DNAs and stimulated the cells with a focal and transient electrical stimulus that promotes the release of exosomes carrying transcribed mRNAs and targeting peptides. Compared with bulk electroporation and other exosome-production strategies, cellular nanoporation produced up to 50-fold more exosomes and a more than 103-fold increase in exosomal mRNA transcripts, even from cells with low basal levels of exosome secretion. In orthotopic phosphatase and tensin homologue (PTEN)-deficient glioma mouse models, mRNA-containing exosomes restored tumour-suppressor function, enhanced inhibition of tumour growth and increased survival. Cellular nanoporation may enable the use of exosomes as a universal nucleic-acid carrier for applications requiring transcriptional manipulation.

/pdf/large-scale-generation-of-functional-mrna-encapsulating-4psl4mqibx.pdf

Large-scale generation of functional mRNA-encapsulating exosomes via cellular nanoporation.

Majority of the human genome is transcribed to RNAs that do not encode proteins. These non-coding RNAs (ncRNAs) play crucial roles in regulating the initiation and progression of various cancers. Given the importance of the ncRNAs, the roles of ncRNAs in cancers have been reviewed elsewhere. Thus, in this review, we mainly focus on the recent studies of the function, regulatory mechanism and therapeutic potential of the ncRNAs including microRNA (miRNA), long ncRNA (lncRNA), circular RNA (circRNA) and PIWI interacting RNA (piRNA), in different type of cancers.

/pdf/non-coding-rna-in-cancer-4r5yke1gns.pdf

Non-coding RNA in cancer.

Tumor cells expressing programmed cell death ligand 1 (PD-L1) interact with PD-1 on CD8+ cytotoxic T lymphocytes (CTLs) to inhibit CTL effector function. In gastric cancer, the mechanism regulating PD-L1 is unclear. The Hedgehog (Hh) signaling pathway is reactivated in various cancers including gastric. Here we tested the hypothesis that Hh-induced PD-L1 inactivates effector T cell function and allows gastric cancer cell proliferation. Mouse organoids were generated from tumors of a triple-transgenic mouse model engineered to express an activated GLI2 allele, GLI2A, in Lgr5-expressing stem cells, (mTGOs) or normal mouse stomachs (mGOs). Bone marrow-derived dendritic cells (DCs) were pulsed with conditioned media collected from normal (mGOCM) or cancer (mTGOCM) organoids. Pulsed DCs and CTLs were then co-cultured with either mGOs or mTGOs in the presence of PD-L1 neutralizing antibody (PD-L1Ab). Human-derived gastric cancer organoids (huTGOs) were used in drug and xenograft assays. Hh/Gli inhibitor, GANT-61 significantly reduced the expression of PD-L1 and tumor cell proliferation both in vivo and in vitro. PD-L1Ab treatment induced tumor cell apoptosis in mTGO/immune cell co-cultures. GANT-61 treatment sensitized huTGOs to standard-of-care chemotherapeutic drugs both in vivo and in vitro. Thus, Hh signaling mediates PD-L1 expression in gastric cancer cells and subsequently promotes tumor proliferation.

/pdf/hedgehog-signaling-induces-pd-l1-expression-and-tumor-cell-249utahfli.pdf

Hedgehog signaling induces PD-L1 expression and tumor cell proliferation in gastric cancer.

MicroRNAs (miRNAs) are short, non-coding, conserved, oligonucleotides that are regulatory in nature and are often dysregulated in many cancers including prostate cancer. Depending on the level of complementarity between the miRNA and mRNA target, they can either inhibit translation or degrade the target mRNA. MiRNAs expression is specific to the type of cancer, its stage and level of metastasis, making miRNAs potential stage-specific biomarkers of cancer. Recent research has shown that these miRNAs have the potential to be a diagnostic and prognostic non-invasive biomarker for various cancers including prostate cancer. Various miRNAs have been reported as novel biomarkers for prostate cancer therapy. However, there is inconsistency in the data reported and no overlapping expression pattern could be found. In this review, we have highlighted the most consistently reported dysregulated miRNAs in prostate cancer from the existing literature and discussed the currently available data on their role in regulating the hallmarks of prostate cancer. These four most consistently reported dysregulated miRNAs viz. miRNA-141, miRNA-375, miRNA-221 and miRNA-21 need to be further validated in terms of their regulatory potential in regulating various pathways important for prostate cancer management.

The microRNA signatures: aberrantly expressed miRNAs in prostate cancer.

MicroRNA biogenesis, gene silencing mechanisms and role in breast, ovarian and prostate cancer.

MiR-141-3p promotes prostate cancer cell proliferation through inhibiting kruppel-like factor-9 expression.

Background
The aim of this study was to establish a culture method for mouse dendritic cells (DCs) in vitro and observe their morphology at different growth stages and their ability to induce the proliferation of T lymphocytes.

/pdf/culture-and-identification-of-mouse-bone-marrow-derived-1w09chb1g8.pdf

Culture and Identification of Mouse Bone Marrow-Derived Dendritic Cells and Their Capability to Induce T Lymphocyte Proliferation.

ABSTRACT Interstitial Cells of Cajal (ICC) plays a critical role in the peristaltic contractions of the gastrointestinal and urinary tract. The dysfunction and loss of ICC contributes to hypokinetic disease, such as gallstoneand ureteropelvic junction obstruction . In the present study, we identified the underlying driving molecular signals of oxidative stress and apoptosis in ICC. ICC was isolated from small intestine of Balb/c mice, and stimulated with tumor necrosis factor-alpha (TNF-α). MTT and flow cytometry were performed to assess cell viability, apoptosis, and the level of reactive oxygen species in ICC, respectively. The level of malondialdehyde, superoxide dismutase, and glutathione peroxidase in cells were measured to assess oxidative stress. The expression of inflammatory factors (interleukin, IL-1 and IL-6) and apoptosis-related proteins were detected by western blot. We observed that TNF-αinduced inflammation, oxidative stress and cell apoptosis in ICC. By using quantitative real-time PCR , we verified that the expression of long non-coding RNAMEG3 was elevated by TNF-α in ICC. Silencing MEG3 reversed inflammation, oxidative stress, and cell apoptosisin TNF-α-treated ICC. Subsequently, we confirmed that MEG3 sponged cytoprotective miR-21 to upregulate the expression of I-kappa-B-kinase beta (IKKB) and activate the nuclear factor kappa-B (NF-κB) pathway. Both miR-21 overexpression and IKKB knockdown reduced TNF-α-induced above symptoms in ICC. Taken together, we can conclude that MEG3 mediates inflammation, oxidative stress and apoptosis in TNF-α-treated ICC via the miR-21/IKKB-NF-κB axis. The study improves our understanding of the molecular mechanism of ICC reduction related diseases. Graphical Abstract

https://www.tandfonline.com/doi/pdf/10.1080/21655979.2022.2054501?needAccess=true

Long non-coding RNA MEG3 promotes tumor necrosis factor-alpha induced oxidative stress and apoptosis in interstitial cells of cajal via targeting the microRNA-21 /I-kappa-B-kinase beta axis

The aim of this work was to create a xenogeneic cell scaffold complex with rabbit bladder acellular matrix and rat hair follicle stem cells, to study the feasibility of construct tissue engineer bladder through biocompatibility of hair follicle stem cells and heterogeneous bladder acellular matrix. New Zealand rabbit bladder acellular matrix was prepared. Scanning electron microscope and Masson staining were used to analyse the acellular material. Two-steps precipitation method was used to place the third generation of hair follicle stem cells onto the surface of the bladder acellular matrix. The in vitro cell growth on the scaffold complex was regularly monitored through an inverted microscope. Cell growth curve was established and histological examination and scanning electron microscopic were used to analyse the progresses of the cell growth on the matrix material. The prepared bladder acellular matrix was white, translucent and membranous. It possessed a fibrous network and collagen structure without any significant cell residues as displayed by the scanning electron microscope, and Masson staining. After 48 h of culture, observation by inverted microscope showed that the hair follicle stem cells grew well around the bladder acellular matrix. After 1 week of culture, scanning electron microscopy showed that the hair follicle stem cells spread and adhered on the surface of the scaffold. The in vitro culture of rat hair follicle stem cells and the rabbit bladder acellular matrix possessed a good biocompatibility, which provides a good experiment support for hair follicle stem cells to repair the bladder defects disease.

/pdf/in-vitro-culture-of-rat-hair-follicle-stem-cells-on-rabbit-1c0d7ybntb.pdf

Jia Li

Papers

MiR-141-3p promotes prostate cancer cell proliferation through inhibiting kruppel-like factor-9 expression.

Culture and Identification of Mouse Bone Marrow-Derived Dendritic Cells and Their Capability to Induce T Lymphocyte Proliferation.

Long non-coding RNA MEG3 promotes tumor necrosis factor-alpha induced oxidative stress and apoptosis in interstitial cells of cajal via targeting the microRNA-21 /I-kappa-B-kinase beta axis

In vitro culture of rat hair follicle stem cells on rabbit bladder acellular matrix.

The histocompatibility research of hair follicle stem cells with bladder acellular matrix.