Background: Alveolar type 2 (AT2) cells and bronchioalveolar stem cells (BASC) perform critical regenerative functions in response to lung damage. Published data show that nonhematopoietic, bone marrow-derived “very small embryonic-like stem cells” (VSELs) can differentiate in vivo into surfactant protein C (SPC)-producing AT2 cells in the lung. Here, we test directly whether VSEL-derived BASC and AT2 cells function to produce differentiated progeny. Methods: using a reporter mouse in which the H2B-GFP fusion protein is driven from the murine SPC promoter, we tested whether bone marrow-derived VSELs or non-VSEL/nonhematopoietic stem cells (non-VSEL/non-HSCs) can differentiate into AT2 and BASC cells that function as progenitor cells. Immediately following bleomycin administration, WT recipient mice underwent intravenous administration of VSELs or non-VSEL/non-HSCs from SPC H2B-GFP mice. GFP+ AT2 and BASC were isolated and tested for progenitor activity using in vitro organoid assays. Results: after 21 days in vivo, we observed differentiation of VSELs but not non-VSEL/non-HSCs into phenotypic AT2 and BASC consistent with previous data in irradiated recipients. Subsequent in vitro organoid assays revealed that VSEL-derived AT2 and BASC maintained physiological potential for differentiation and self-renewal. Conclusion: these findings prove that VSELs produce functional BASC and AT2 cells, and this may open new avenues using VSELs to develop effective cell therapy approaches for patients with lung injury.

The effect of a sucrose diet and repeated one-day starvation on oxidative status in the ovary and uterus is still unknown. Our analysis focused on carbohydrate-lipid metabolism parameters and the changes in red blood cells, ovary and uterus superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) activities and malonylodialdehyde (MDA) concentration in rats fed with a diet containing 16% of sucrose and subjected to systematic one-day starvation when using such a diet. It was found that a diet with 16% sucrose contributed to the increase of antioxidant enzyme activity in the blood (GPx and CAT) and uterus (SOD), without changes in MDA concentrations, which indicates an increase in reactive oxygen species (ROS) concentration in these tissues, being balanced by an increase in antioxidant enzyme activity. The introduction of a regular one-day starvation period into the diet intensified oxidative stress and led to a redox imbalance in the reproductive tissues of female rats. This was manifested by higher GPx activity, lower CAT activity and higher MDA concentration in the uterus and lower GPx and CAT activities and lower MDA concentration in the ovaries. The observed changes may be the cause of fertility disorders and possible problems with fertilised egg cell implantation into the uterine tissue.

Background: Bone marrow-derived mononuclear cells (BM-MNC) consist of a heterogeneous mix of mesenchymal stem cells (MSC), hematopoietic progenitor cells (HPC), endothelial progenitor cells (EPC), monocytes, lymphocytes and pluripotent stem cells. Whereas the importance of MSC and EPC has been well documented in bone healing and regeneration studies, the role of pluripotent stem cells is still poorly understood. In the present study we evaluated if and how Very Small Embryonic Like cells (VSEL), isolated from rat BM-MNC, contribute to bone healing. Methods: Large bone defects were made in the femurs of 38 Sprague Dawley female rats and treated with β-TCP scaffold granules seeded with male VSEL; BM-MNC, VSEL-depleted BM-MNC or scaffold alone, and bone healing was evaluated at 8 weeks post-surgery. Results: Bone healing was significantly increased in defects treated with VSEL and BM-MNC, compared to defects treated with VSEL-depleted BM-MNC. Donor cells were detected in new bone tissue, in all the defects treated with cells, and in fibrous tissue only in defects treated with VSEL-depleted BM-MNC. The number of CD68+ cells was the highest in the VSEL-depleted group, whereas the number of TRAP positive cells was the lowest in this group. Conclusions: Based on the results, we can conclude that VSEL play a role in BM-MNC induced bone formation. In our rat femur defect model, in defects treated with VSEL-depleted BM-MNC, osteoclastogenesis and bone formation were decreased, and foreign body reaction was increased.

Dehydroepiandrosterone (DHEA) is an abundant steroid and precursor of sex hormones. During aging, the reduction in DHEA synthesis causes a significant depletion of estrogens and androgens in different organs, such as the ovaries, brain, and liver. Primary Biliary Cholangitis (PBC) is a cholestatic liver disease that begins with immune-mediated bile duct damage, and is followed by liver fibrosis, and finally, cirrhosis. PBC primarily affects postmenopausal women, with an average age of diagnosis of 65 years, but younger women are also affected. Here, we analyzed the levels of DHEA, estradiol (E2), and estriol (E3) in the PBC sera of females at an age of diagnosis under 40 (n = 37) and above 65 (n = 29). Our results indicate that in PBC patients at an age of diagnosis under 40, E2 was significantly lower compared to that in healthy women. In contrast, the levels of DHEA and E3 were in a normal range. Furthermore, ELISA assays revealed that in PBC patients at an age of diagnosis above 65, the levels of DHEA, E2, and E3 significantly declined in comparison to those in younger patients. In addition, flow cytometry analysis showed that the level of IL-8 significantly decreased while the level of TNF-α increased in older PBC patients compared to younger ones. Moreover, we showed for the first time that the sulfonated form of DHEA, DHEA-S, reduces the levels of both pro-inflammatory interleukins, IL-8 and TNF-α, in PBC-like cholangiocytes (H69-miR506), while it diminishes the level of the pro-fibrotic interleukin, IL-13, in hepatocytes (Hep-G2). Finally, we demonstrated that the expression of the pro-fibrotic agent TGF-β significantly increased in both the early (F0–F3) and cirrhotic (F4) stages of PBC, and this elevation was accompanied by higher α-SMA expression.

Very small embryonic-like stem cells (VSELs) are a dormant population of development early stem cells deposited in adult tissues that as demonstrated contribute to tissue/organ repair and regeneration. We postulated developmental relationship of these cells to migrating primordial germ cells (PGCs) and explained the quiescent state of these cells by the erasure of differently methylated regions (DMRs) at some of the paternally imprinted genes involved in embryogenesis. Recently, we reported that VSELs began to proliferate and expand in vivo in murine bone marrow (BM) after exposure to nicotinamide (NAM) and selected pituitary and gonadal sex hormones. In the current report, we performed proteomic analysis of VSELs purified from murine bone marrow (BM) after repeated injections of NAM + Follicle-Stimulating Hormone (FSH) that in our previous studies turned out to be an effective combination to expand these cells. By employing the Gene Ontology (GO) resources, we have performed a combination of standard GO annotations (GO-CAM) to produce a network between BM steady-state conditions VSELs (SSC-VSELS) and FSH + NAM expanded VSELs (FSH + NAM VSELs). We have identified several GO biological processes regulating development, organogenesis, gene expression, signal transduction, Wnt signaling, insulin signaling, cytoskeleton organization, cell adhesion, inhibiting apoptosis, responses to extra- and intracellular stimuli, protein transport and stabilization, protein phosphorylation and ubiquitination, DNA repair, immune response, and regulation of circadian rhythm. We report that VSELs express a unique panel of proteins that only partially overlapped with the proteome of BM – derived hematopoietic stem cells (HSCs) and hematopoietic mononuclear cells (MNCs) and respond to FSH + NAM stimulation by expressing proteins involved in the development of all three germ layers. Thus, our current data supports further germ-lineage origin and multi germ layer differentiation potential of these cells. Graphical abstract

The flow cytometry method (FCM) is a widely renowned practice increasingly used to assess the microbial viability of probiotic products. Additionally, the measurement of water activity (a_w) can be used to confirm the presence of viable cells in probiotic products throughout their shelf lives. The aim of this study was to investigate the correlation between changes in a_w and variations in active fluorescent units (AFU), a unit commonly used in flow cytometry method, during the aging of probiotic products containing freeze-dried bacteria. We controlled the stability of probiotic products for bacterial counts (using ISO 19344 method) and a_w levels in commercially available capsules containing freeze-dried bacteria such as Lactobacillus sp. or combinations of Lactobacillus sp. and Bifidobacterium sp. in standard conditions (25 ± 2 °C and 60% relative humidity) over a period of 24 months. During this time, the bacterial contents decreased by 0.12 Log₁₀ in the single-strain product, by 0.16 Log₁₀ in the two-strain product and by 0.26 Log₁₀ in the multi-strain product. With the increase in a_w, the number of bacteria decreased but the a_w at the end point of the stability study did not exceed 0.15 in each of the three tested products. FCM combined with a_w is a prospective analysis that can be used to assess the stability of probiotic products, both for its ability to detect bacterial viability and for practical (analysis time) and economic reasons.