LncRNA XIST promotes extracellular matrix synthesis, proliferation and migration by targeting miR-29b-3p/COL1A1 in human skin fibroblasts after thermal injury.

BACKGROUND: Long noncoding RNAs (lncRNAs) have been reported to be associated with dermis process during burn wound healing. This study aimed to investigate the role of lncRNA X-inactive specific transcript (XIST) in human skin fibroblasts (HSF) and extracellular matrix (ECM) as well as the regulatory network of XIST/microRNA-29b-3p (miR-29b-3p)/collagen 1 alpha 1 (COL1A1). METHODS: The wound samples were collected from 25 patients with deep partial thickness burn at day 5 after burn. The thermal injured model was established using HSF cells. The expressions of XIST, miR-29b-3p and COL1A1 were measured by quantitative real-time polymerase chain reaction and western blot. ECM synthesis, cell proliferation and migration were detected by western blot, cell counting kit-8 and trans-well assays, respectively. The interaction between miR-29b-3p and XIST or COL1A1 was explored by bioinformatics analysis and luciferase reporter assay. RESULTS: The expressions of XIST and COL1A1 were enhanced but miR-29b-3p expression was decreased after thermal injury. XIST overexpression promoted ECM synthesis, cell proliferation and migration in thermal injured HSF cells. However, XIST knockdown played an opposite effect. miR-29b-3p overexpression inhibited ECM synthesis, cell proliferation and migration, which was reversed by XIST. COL1A1 silence suppressed ECM synthesis, cell proliferation and migration by miR-29b-3p targeting. Moreover, COL1A1 up-regulation weakened the effect of XIST silence on ECM synthesis and HSF cell function. CONCLUSION: XIST promoted ECM synthesis, cell proliferation and migration by sponging miR-29b-3p and targeting COL1A1 in HSF cells after thermal injury, indicating the promoting role of XIST in wound healing.

Overexpression of salt-induced protein (salT) delays leaf senescence in rice.

Senescence, a highly programmed process, largely determines yield and quality of crops. However, knowledge about the onset and progression of leaf senescence in crop plants is still limited. Here, we report that salt-induced protein (salT), a new gene, may be involved in leaf senescence. Overexpressing salT could prolong the duration of leaves with higher concentrations of chlorophyll compared with the wild type. Moreover, overexpression of salT could delay the senescence of rice leaves though the inhibition of senescence associated genes (SAGs). Overall, the characterization of salT suggested that it is a new gene affecting the leaf senescence induced by natural and dark conditions.

LncRNA XIST promotes extracellular matrix synthesis, proliferation and migration by targeting miR-29b-3p/COL1A1 in human skin fibroblasts after thermal injury

Background: Long noncoding RNAs (lncRNAs) have been reported to be associated with dermis process during burn wound healing. This study aimed to investigate the role of lncRNA X-inactive specific transcript (XIST) in human skin fibroblasts (HSF) and extracellular matrix (ECM) as well as the regulatory network of XIST/microRNA-29b-3p (miR-29b-3p)/collagen 1 alpha 1 (COL1A1). Methods: The wound samples were collected from 25 patients with deep partial thickness burn at day 5 after burn. The thermal injured model was established using HSF cells. The expressions of XIST, miR-29b-3p and COL1A1 were measured by quantitative real-time polymerase chain reaction and western blot. ECM synthesis, cell proliferation and migration were detected by western blot, cell counting kit-8 and trans-well assays, respectively. The interaction between miR-29b-3p and XIST or COL1A1 was explored by bioinformatics analysis and luciferase reporter assay. Results: The expressions of XIST and COL1A1 were enhanced but miR-29b-3p expression was decreased after thermal injury. XIST overexpression promoted ECM synthesis, cell proliferation and migration in thermal injured HSF cells. However, XIST knockdown played an opposite effect. miR-29b-3p overexpression inhibited ECM synthesis, cell proliferation and migration, which was reversed by XIST. COL1A1 silence suppressed ECM synthesis, cell proliferation and migration by miR-29b-3p targeting. Moreover, COL1A1 up-regulation weakened the effect of XIST silence on ECM synthesis and HSF cell function. Conclusion: XIST promoted ECM synthesis, cell proliferation and migration by sponging miR-29b-3p and targeting COL1A1 in HSF cells after thermal injury, indicating the promoting role of XIST in wound healing.

The complete mitochondrial genome of the Cavia aperea.

Cavia aperea which is a Brazilian guinea pig is found in the South America. Recently the genome sequencing of C. aperea was done, but no more information of its mitochondrial had been reported. Herein, we assembled the complete mitochondrial genome sequence of C. aperea. It is a 16 835 bp long sequence with most mitogenome's characteristic structure; 13 protein-coding genes, 2 rRNA genes, 22 tRNA genes, 1 D-loop region, 1 repeat region and 3 STS regions. The GC-content of our fresh sequence is 39%. It can verify the accuracy and utility of newly determined mitogenome sequences by the phylogenetic analysis, based on whole mitogenome alignment with C. porcellus, which is the closest relative to C. aperea. We expect that using the full mitogenome we can address the taxonomic issues and study the related the evolution events.

Co-culture of apoptotic breast cancer cells with immature dendritic cells: a novel approach for DC-based vaccination in breast cancer

A dendritic cell (DC)-based vaccine strategy could reduce the risk of recurrence and improve the survival of breast cancer patients. However, while therapy-induced apoptosis of hepatocellular and colorectal carcinoma cells can enhance maturation and antigen presentation of DCs, whether this effect occurs in breast cancer is currently unknown. In the present study, we investigated the effect of doxorubicin (ADM)-induced apoptotic MCF-7 breast cancer cells on the activation of DCs. ADM-induced apoptotic MCF-7 cells could effectively induce immature DC (iDC) maturation. The mean fluorescence intensity (MFI) of DC maturity marker CD83 was 23.3 in the ADM-induced apoptotic MCF-7 cell group compared with 8.5 in the MCF-7 cell group. The MFI of DC co-stimulatory marker CD86 and HLA-DR were also increased after iDCs were treated with ADM-induced apoptotic MCF-7 cells. Furthermore, the proliferating autologous T-lymphocytes increased from 14.2 to 40.3% after incubated with DCs induced by apoptotic MCF-7 cells. The secretion of interferon-γ by these T-lymphocytes was also increased. In addition, cell-cell interaction between apoptotic MCF-7 cells and iDCs, but not soluble factors released by apoptotic MCF-7 cells, was crucial for the maturation of iDCs. These findings constitute a novel in vitro DC-based vaccine strategy for the treatment of breast cancer by ADM-induced apoptotic MCF-7 cells.

Co-culture of apoptotic breast cancer cells with immature dendritic cells: a novel approach for DC-based vaccination in breast cancer.

A dendritic cell (DC)-based vaccine strategy could reduce the risk of recurrence and improve the survival of breast cancer patients. However, while therapy-induced apoptosis of hepatocellular and colorectal carcinoma cells can enhance maturation and antigen presentation of DCs, whether this effect occurs in breast cancer is currently unknown. In the present study, we investigated the effect of doxorubicin (ADM)-induced apoptotic MCF-7 breast cancer cells on the activation of DCs. ADM-induced apoptotic MCF-7 cells could effectively induce immature DC (iDC) maturation. The mean fluorescence intensity (MFI) of DC maturity marker CD83 was 23.3 in the ADM-induced apoptotic MCF-7 cell group compared with 8.5 in the MCF-7 cell group. The MFI of DC co-stimulatory marker CD86 and HLA-DR were also increased after iDCs were treated with ADM-induced apoptotic MCF-7 cells. Furthermore, the proliferating autologous T-lymphocytes increased from 14.2 to 40.3% after incubated with DCs induced by apoptotic MCF-7 cells. The secretion of interferon-γ by these T-lymphocytes was also increased. In addition, cell-cell interaction between apoptotic MCF-7 cells and iDCs, but not soluble factors released by apoptotic MCF-7 cells, was crucial for the maturation of iDCs. These findings constitute a novel in vitro DC-based vaccine strategy for the treatment of breast cancer by ADM-induced apoptotic MCF-7 cells.

Comparison of expression of monomeric and multimeric adenoregulin genes in Escherichia coli and Pichia pastorias.

Adenoregulin is a 33 amino acid antibiotic peptide who belongs to dermaseptin family which is the first vertebrate family to show lethal effects against filamentous fungi, as well as a broad spectrum of pathogenic microorganisms. Synthetic adenoregulin gene was cloned in 2, 4 and 6 tandem repeats and subcloned in pET32a and pET22b vectors. Recombinant plasmids were transformed into E. coli BL21(DE3), Fusion proteins of Trx-ADR1, Trx-ADR2 and Trx-ADR4 could be expressed after the hosts were induced by IPTG, but the expression level decreased dramatically with the number of tandem repeats increased. ADR1, ADR4 and ADR6 could not be expressed by E. coli without carrier proteins. But for Pichia pastoris GS115, ADR1 and ADR6 in the fermentation broth of the hosts could be detected by ELISA, and the bactericidal activities could also be observed.

Technetium-99m labeled antisense oligonucleotide-noninvasive tumor imaging in mice

The 15-mer oligonucleotide sequence was synthesized, aminolinked (sense and antisense phosphodiester) and conjugated with S-Acetyl-NHS-MAG3 by a N-hydroxy-succinimide derivative. The purified MAG3-DNA was radiolabeled with 99mTc by transchelation from sodium tartrate and free 99mTc was separated by gel filtration. The radiolabeled antisense and sense probes were injected intravenously in mammary tumor-bearing KM mice(1×106 cells,6 days post inoculation).Biodistribution was studied and the mice were imaged.Essentially complete conjugation was achieved by reverse-phase Sep-Pak C18 chromatography analysis.The MAG3-DNA was labeled with 99mTc at room temperature and neutral pH, with a mean labeling efficiency of 80.11 percent (s.d=2.96 percent , N=4). After labeling, the stability of the DNA in saline or serum was retained as determined by reverse-phase Sep-Pak C18 chromatography analysis, except a shift at 30 min in serum incubation which might suggest a short time serum protein binding. The labeled antisense DNA still remained the ability to hybridize with its complementary DNA. The pharmacodynamics of 99mTc labeled c-myc probes (antisense and sense) in mammary tumor-bearing KM mice did not change with the time postinjection. The highest accumulation of label was in the liver first, with the kidney and small bowel next. The injected activity localized in the lesion as early as 30min and reached a saturation value at 4hr. The accumulation of radioactivity in the tumor was lower at all time points in animals receiving the blocking oligonucleotides or sense probes. All images obtained with 99mTc-MAG3-c-myc antisense probes showed specific accumulation of radioactivity at the site of tumor. Radiolabel rapidly accumulates at the site of tumor and remains associated with the site even though circulation levels of radioactivity have greatly diminished. The tumor was readily evident since 45min and reached the highest tumor-to-muscle ratio at 4hr. The quite encouraging result was obtained at 20hr to 22hr when the background activity was diminished sufficiently. Positive imaging was not obtained in case of control group (in which non-conjugated, non-labeled antisense oligonucleotides were administered 2hr before the radiolabeled antisense probes were injected) and of sense group. Conclusion The 99mTc labeled antisense probe may provide a sensible and specific tool for noninvasive imaging of c-myc oncogene mRNA for a variety of malignant tumors at an earlier stage