Microfluidic synthesis of retinoic acid-loaded nanoparticles for neural differentiation of trabecular meshwork mesenchymal stem cell.

OBJECTIVES: This study aimed to fabricate the PCL-nanoparticles (NPs) loaded retinoic acid (RA) using the microfluidic system for successful cellular uptake and induction of neuronal differentiation of trabecular meshwork mesenchymal stem cells (TMMSCs). METHODS: A microfluidic system used to synthesize RA-loaded NPs, DLS, FTIR, TEM, and UV-spectroscopy was recruited to characterize and study the release of RA. Also, the toxicity, cellular uptake, and neuronal differential of TMMSCs have been assessed. RESULTS: According to the obtained results, the spherical NPs (117.6±0.35 nm, ‒19.4±5.3) and RA-loaded NPs (121.6±0.75 nm, ‒23.6±1.3) were synthesized successfully by microfluidic system. 7.8±2.04 % of RA was loaded in NPs, and 25 % was released in the first four hours. Thus, the NPs have been successfully internalized into the stem cells, leading to a significant increase in neural genes and protein (ß Tubulin III and Map-2) expression. CONCLUSION: Our study's harvested results have represented valid data for practical use of microfluidic systems in the term of NPs loaded RA synthesis and its successful function to cellular internalization and euronal differentiation of TMMSCs (Tab. 2, Fig. 10, Ref. 46).

TRAIL-expressing recombinant Lactococcus lactis induces apoptosis in human colon adenocarcinoma SW480 and HCT116 cells.

AIMS: We investigated the ability of Lactococcus lactis, a species generally regarded as safe, to express tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) protein. The expressed protein was either cell wall anchored or secreted, and it was assessed whether this could induce apoptosis in human colon adenocarcinoma cell lines SW480 and HCT116. METHODS AND RESULTS: Constructs were designed to produce either secreted or cell wall-anchored forms of human TRAIL cloned into pNZ7021 expression vector. The expression by L. lactis was confirmed by Western blotting and immunofluorescence. Induction of cell death was evaluated by coculturing transformants producing either form of TRAIL protein with the two cell lines followed by MTT assay. Gene expression of apoptosis genes, Bax and Bcl2, was assessed by qPCR. The viability of SW480 and HCT116 cells treated with recombinant L. lactis was significantly reduced. A significant change was observed in the ratio of Bax/Bcl2 expression in HCT116 cells only following treatment with the supernatant of recombinant L. lactis containing secreted TRAIL. CONCLUSION: Recombinant L. lactis producing TRAIL protein can induce apoptosis in human colon adenocarcinoma cell lines SW480 and HCT116. SIGNIFICANCE AND IMPACT OF THE STUDY: The use of recombinant probiotics that produce anticancer compounds is a promising option for combating cancer cells.

A review on prolificacy genes in sheep.

Ovulation rate and litter size are important reproduction traits in sheep and are of high economic value. Reproduction traits typically have low to medium heritabilities and do not exhibit a noticeable response to phenotypic selection. Therefore, inclusion of genetic information of the genes associated with reproductive ability could efficiently enhance the selection response. The most important major genes affecting prolificacy and their genetic diversities in different sheep breeds were reviewed. Different causative mutations with major effects on reproductive traits including ovulation rate and litter size have been found in various sheep breeds around the world. A general overview of the studies on main prolificacy genes showed that some alleles may express different phenotypic effects in different breeds, and thus, further studies on epistatic effects are necessary for more understanding of genetic control of reproductivity in sheep. Regarding the polygenic control of fertility traits, application of new high-throughput technologies to find new variants is essential for future studies. Moreover, genomewide association studies and genomic best linear unbiased predictions of breeding values are likely to be effective tools for genetic improvement of sheep reproductive performance traits.

Novel Mutation in Exon 1 of the BMP15 Gene and its Association with Reproduction Traits in Sheep.

The BMP15 gene is a growth factor and a member of the transforming growth factor ß (TGFß) superfamily, specifically expressed in oocytes. In the present study, polymorphism of BMP15 gene exon 1 was studied using single strand conformational polymorphism (SSCP) and direct DNA sequencing methods in 170 Mehraban and Lori sheep ewes. A 231-bp fragment in BMP15 exon 1 was amplified by PCR reactions. Two genotypes (GG and AG) with a new point mutation at position 121 bp of the studied fragment (c.379G>A in reference GenBank number AF236078.1 sequence), deducing an amino acid exchange in the codified amino acid sequence (p.Glu41Lys) were identified in the studied populations. The AG and GG frequencies were 74.4% and 25.6% in Mehraban and 44.7% and 55.3% in Lori sheep, respectively. Frequencies of the A and G alleles were 37.2% and 62.8% in Mehraban and 22.4% and 77.6% in Lori sheep, respectively. Two different secondary structures of protein were predicted for encoded precursor protein. The genotypes GG and AG did not have any significant association with the studied reproductive traits, but the AA genotype is likely to have a lethal or sterility effect.

Comparative analysis of mesenchymal stromal cells from murine bone marrow and amniotic fluid.

BACKGROUND: Murine mesenchymal stromal cells (mMSC) are a good model for pre-clinical investigations, and alternative sources of mMSC are subject to intensive experiments. In the present study, we obtained mMSCs from amniotic fluid (AF) and compared their characteristics with mMSCs from bone marrow (BM). METHODS: NMRI mice, 4-6 weeks old, were killed and AF and BM cells collected from those in the second week of pregnancy. MSC were achieved by adhesion to cell culture plastic. Isolated cells were assessed for clonogenic capacity, some surface markers (epitopes) and differentiation potential. RESULTS: We achieved AF mMSC more readily than BM mMSC. Differences concerning colony assay and some surface markers of mMSC derived from these two source cells were observed. Most strikingly, AF mMSC showed no adipogenic differentiation capacity, in contrast with BM mMSC. DISCUSSION: Our results show that mMSC from AF are an appropriate source for pre-clinical investigation. Furthermore, mMSC from different sources of mice vary in clonogenic capacity, surface markers and differentiation potential.