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The aim of this study was to compare the effectiveness of the Gaussian and Flat-Top profiles in proliferation and differentiation of mesenchymal stem cell of buccal fat pad. Based on the timing schedule and type of laser handpieces, the cells were assigned to a control group with no radiation, and two irradiation test groups (980 nm) with Flat-Top (F) (power of 1.1 W, beam area of 1 cm 2 ) and standard Gaussian (G) (power of 0.7 W, beam area of 0.5 cm 2 ) handpieces. Each test group was divided into three subgroups, receiving one time (60 J/cm 2 ), two times (120 J/cm 2 ), and three times (180 J/cm 2 ) irradiation. 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) and Annexin V tests were performed. The Alizarin Red staining and polymerase chain reaction tests were done both at the beginning and the end of the first and second weeks. The degree of mineralization and expression of osteogenic markers, RUNX2, OCN, and OPN were evaluated. Based on the MTT and Annexin V test results, both test groups outperformed the control group in degrees of cell proliferation during the first day of laser irradiation (p < 0.05). After one and two times irradiation, the expression of osteogenic markers in the test groups was significantly higher than the control group. PBM with Flat-Top and Gaussian handpieces can enhance ossification and cell differentiation regardless of the type of handpieces., (© 2024 American Society for Photobiology.)

Purpose: Previous studies have shown that the mean dose to the parotid gland stem cell rich regions (D mean,SCR ) is the strongest dosimetric predictor for the risk of patient-reported daytime xerostomia. This study aimed to test whether the relationship between patient-reported xerostomia and D mean,SCR is explained by a dose-dependent reduction of saliva production., Methods and Materials: In 570 patients with head and neck cancer treated with definitive radiation therapy (RT), flow from the parotid (FLOW PAR ) and submandibular/sublingual (FLOW SMSL ) glands, and patient-reported daytime (XER DAY ) and nighttime (XER NIGHT ) xerostomia were prospectively measured before, at 6 months, and 12 months after RT. Using linear mixed effect models, the relationship of the mean dose to the parotid glands (D mean,par ), D mean,SCR , non-SCR parotid gland tissue (D mean,non-SCR ), submandibular glands (D mean,sub ), and oral cavity (D mean,oral ) with salivary flow and xerostomia was analyzed while correcting for known confounders., Results: D mean,SCR proved to be responsible for the effect of D mean,par on FLOW PAR (P ≤ .03), while D mean,non-SCR did not affect FLOW PAR (P ≥ .11). To illustrate, increasing D mean,SCR by 10 Gy at a fixed D mean,non-SCR reduced FLOW PAR by 0.02 mL/min (25%) after RT. However, if the opposite happened, no change in FLOW PAR was observed (0.00 mL/min [4%]). As expected, D mean,sub was significantly associated with FLOW SMSL (P < .001). For example, increasing D mean,sub by 10 Gy reduced FLOW SMSL by 0.07 mL/min (26%) after RT. Xerostomia scores were also affected by dose to the salivary glands. D mean,SCR and D mean,oral were associated with higher XER DAY scores (P ≤ .05), while D mean,sub increased XER NIGHT scores (P = .01). For example, an increase of 10 Gy in D mean,SCR raised XER DAY scores by 2.13 points (5%) after RT, while an additional 10 Gy in D mean,subs increased XER NIGHT scores by 2.20 points (6%) after RT. Salivary flow was not only associated with radiation dose, but also with xerostomia scores in line with the salivary glands' functions; ie, FLOW PAR only influenced XER DAY (P < .001, 10.92 points lower XER DAY per 1 mL/min saliva), while FLOW SMSL affected XER DAY and XER NIGHT (P ≤ .004, 6.69 and 5.74 points lower XER DAY and XER NIGHT , respectively, per 1 mL/min saliva). Therefore, the observed relationships between dose and xerostomia were corrected for salivary flow. As hypothesized, D mean,SCR only increased XER DAY scores via reducing FLOW PAR , whereas the effects of D mean,oral on XER DAY and D mean,sub on XER NIGHT were independent of salivary flow., Conclusions: Higher SCR region dose reduced parotid gland saliva production, subsequently resulting in higher daytime xerostomia scores. Consequently, this study supports the clinical implementation of stem cell sparing RT to preserve salivary flow with the aim of reducing the risk of xerostomia., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: The present study aimed to investigate the effects of infrared diode laser irradiation on the proliferation and differentiation capacity of periodontal ligament stem cells (hPDLSCs), which are optimal cell sources for periodontal regeneration., Methods: hPDLSCs were isolated and characterized by flow cytometric analysis of mesenchymal stem cell markers, and their trilineage differentiation capacity was tested. hPDLSCs were then cultured and irradiated with infrared diode laser (970 nm) at a power of 200 mW and a fluence of 4 J/cm 2 for 3 s. MTT assay was performed to assess cellular proliferation. Cell cycle analysis was performed, and the impact of infrared diode laser irradiation on the stemness and osteogenic differentiation potential of hPDLSCs was evaluated via RT‒PCR., Results: Infrared diode laser application enhanced the stemness, viability, proliferation, and differentiation of PDLSCs. Stem cell markers (OCT4, SOX2, and NANOG) were significantly upregulated in hPDLSCs exposed to laser irradiation. There was significant overexpression of RUNX2, ALP, OPN, and OCN on day 14 after laser application., Conclusions: These findings provide valuable insights into the specific applications of infrared diode lasers to effectively regenerate periodontal tissues. The results can aid in the development of precise clinical protocols aimed at enhancing osseointegration and promoting tissue regeneration. Ultimately, the combination of infrared diode laser with hPDLSCs is promising for stimulating periodontal regeneration., (© 2024. The Author(s).)

This study assessed the impact of low-level laser irradiation on the viability and proliferation of human periodontal ligament stem cells (hPDLSCs) cultivated on polylactic acid (PLA) scaffolds. hPDLSCs were obtained, characterized, and grown on the surface of PLA films produced via the solvent casting technique. The study involved two groups: the control group, which was not exposed to radiation, and the laser group, which was irradiated with a diode laser (InGaAIP) with a power of 30 mW, a wavelength of 660 nm, and a single dose of 1 J/cm² emitted continuously. Cell viability was assessed 24 and 48 hours after irradiation using the Alamar blue and Live/Dead assays. Flow cytometry was used to assess cell cycle events, and scanning electron microscopy (SEM) was used to evaluate the interaction between cells and the biomaterial. The results revealed a statistically significant increase in cell metabolic activity in the laser group compared with the control group at 24 hours (p <0.05) and 48 hours (p <0.001), as indicated by the Alamar blue assay. The Live/Dead assay also revealed a greater density of viable cells in the laser group. The cell cycle analysis revealed a significant increase in the number of cells in the proliferative phase (G2/M) in the laser group compared with the control group (p <0.001). The SEM images demonstrated that the irradiated group had a greater concentration of cells while still maintaining their cell shape and projections. This study demonstrated that photobiomodulation can increase the viability and proliferation of periodontal stem cells cultured on PLA scaffolds, suggesting the potential of this protocol for future studies on periodontal tissue engineering., (© 2024. The Author(s), under exclusive licence to Springer-Verlag London Ltd., part of Springer Nature.)

The intestinal tract is a typical radiosensitive tissue, and radiation rectal injury is a severe side effect that limits the prescribed dose in radiotherapy of the abdominal and pelvic region. Understanding the post-irradiation kinetics of Lgr5 -positive stem cells is crucial in comprehending this adverse process. In this study, we utilized Lgr5 - EGFP knock-in mice expressing EGFP and LGR5 antibody fluorescence staining of wild-type mice. At the state of radiation injury, the qPCR analysis showed a significant decrease in the expression level of Lgr5 in the rectal epithelial tissue. The dose-response relationship analysis showed that at low to moderate doses up to 10 gray (Gy), Lgr5 -clustered populations were observed at the base of the crypt, whereas at sublethal doses (20 Gy and 29 Gy), the cells exhibited a dot-like scatter pattern, termed Lgr5 -dotted populations. During recovery, 30 days post-irradiation, Lgr5 -clustered populations gradually re-emerged while Lgr5 -dotted populations declined, implying that some of the Lgr5 -dotted stem cell populations re-clustered, aiding regenerations. Based on statistical analysis of the dose-response relationship using wild-type mice, the threshold dose for destroying these stem cell structures is 18 Gy. These findings may help set doses in mouse abdominal irradiation experiments for radiation intestinal injury and for understanding the histological process of injury development.

Radiotherapy is a pivotal intervention for cancer patients, significantly impacting their treatment outcomes and survival prospects. Nevertheless, in the course of treating those with abdominal, pelvic, or retroperitoneal malignant tumors, the procedure inadvertently exposes adjacent intestinal tissues to radiation, posing risks of radiation-induced enteropathy upon reaching threshold doses. Stem cells within the intestinal crypts, through their controlled proliferation and differentiation, support the critical functions of the intestinal epithelium, ensuring efficient nutrient absorption while upholding its protective barrier properties. Intestinal stem cells (ISCs) regulation is intricately orchestrated by diverse signaling pathways, among which are the WNT, BMP, NOTCH, EGF, Hippo, Hedgehog and NF-κB, each contributing to the complex control of these cells' behavior. Complementing these pathways are additional regulators such as nutrient metabolic states, and the intestinal microbiota, all of which contribute to the fine-tuning of ISCs behavior in the intestinal crypts. It is the harmonious interplay among these signaling cascades and modulating elements that preserves the homeostasis of intestinal epithelial cells (IECs), thereby ensuring the gut's overall health and function. This review delves into the molecular underpinnings of how stem cells respond in the context of radiation enteropathy, aiming to illuminate potential biological targets for therapeutic intervention. Furthermore, we have compiled a summary of several current treatment methodologies. By unraveling these mechanisms and treatment methods, we aspire to furnish a roadmap for the development of novel therapeutics, advancing our capabilities in mitigating radiation-induced intestinal damage., (© 2024. The Author(s).)

Background: Vasoactive intestinal peptide (VIP) is a neuronal peptide with prominent distribution along the enteric nervous system. While effects of VIP on intestinal motility, mucosal vasodilation, secretion, and mucosal immune cell function are well-studied, the direct impact of VIP on intestinal epithelial cell turnover and differentiation remains less understood. Intestinal stem and progenitor cells are essential for the maintenance of intestinal homeostasis and regeneration, and their functions can be modulated by factors of the stem cell niche, including neuronal mediators. Here, we investigated the role of VIP in regulating intestinal epithelial homeostasis and regeneration following irradiation-induced injury., Methods: Jejunal organoids were derived from male and female C57Bl6/J, Lgr5-EGFP-IRES-CreER T2 or Lgr5-EGFP-IRES-CreER T2 /R26R-LSL-TdTomato mice and treated with VIP prior to analysis. Injury conditions were induced by exposing organoids to 6 Gy of irradiation (IR). To investigate protective effects of VIP in vivo, mice received 12 Gy of abdominal IR followed by intraperitoneal injections of VIP., Results: We observed that VIP promotes epithelial differentiation towards a secretory phenotype predominantly via the p38 MAPK pathway. Moreover, VIP prominently modulated epithelial proliferation as well as the number and proliferative activity of Lgr5-EGFP +  progenitor cells under homeostatic conditions. In the context of acute irradiation injury in vitro, we observed that IR injury renders Lgr5-EGFP +  progenitor cells more susceptible to VIP-induced modulations, which coincided with the strong promotion of epithelial regeneration by VIP. Finally, the observed effects translate into an in vivo model of abdominal irradiation, where VIP showed to prominently mitigate radiation-induced injury., Conclusions: VIP prominently governs intestinal homeostasis by regulating epithelial progenitor cell proliferation and differentiation and promotes intestinal regeneration following acute irradiation injury., (© 2024. The Author(s).)

Hair loss is a prevalent issue worldwide, which, though not life-threatening, can result in psychological problems, low self-esteem, and social anxiety. Previous studies have shown that ultraviolet radiation can have negative effects on hair follicle cells, leading to hair loss, while the impact of blue light on hair and hair follicle has largely been overlooked. This study aimed to examine the effects of blue light on hair follicle stem cells (HFSCs) and primary dermal papilla cells (DPCs), which are essential components of hair follicles. Human HFSCs and primary DPCs were exposed to blue light (457 nm) at various intensities (1, 4, 8, and 16 mW/cm 2 ) for 3 days. Subsequently, cell viability, cell proliferation, and intracellular reactive oxygen species (ROS) were assessed. The results showed that blue light (457 nm) significantly reduced the cell viability and proliferation of HFSCs and DPCs in vitro, with the inhibition being intensity-dependent. Additionally, blue light triggered the overproduction of ROS in the DPCs. While the exact mechanisms by which blue light affects hair follicle cells remain unclear, these findings suggest that blue light could impede the growth of these cells. This insight may offer a new approach to protecting hair by avoiding exposure to high-intensity blue light., (© 2024. The Author(s), under exclusive licence to Springer-Verlag London Ltd., part of Springer Nature.)

In mice, intestinal tuft cells have been described as a long-lived, postmitotic cell type. Two distinct subsets have been identified: tuft-1 and tuft-2 (ref. 1 ). By combining analysis of primary human intestinal resection material and intestinal organoids, we identify four distinct human tuft cell states, two of which overlap with their murine counterparts. We show that tuft cell development depends on the presence of Wnt ligands, and that tuft cell numbers rapidly increase on interleukin-4 (IL-4) and IL-13 exposure, as reported previously in mice 2-4 . This occurs through proliferation of pre-existing tuft cells, rather than through increased de novo generation from stem cells. Indeed, proliferative tuft cells occur in vivo both in fetal and in adult human intestine. Single mature proliferating tuft cells can form organoids that contain all intestinal epithelial cell types. Unlike stem and progenitor cells, human tuft cells survive irradiation damage and retain the ability to generate all other epithelial cell types. Accordingly, organoids engineered to lack tuft cells fail to recover from radiation-induced damage. Thus, tuft cells represent a damage-induced reserve intestinal stem cell pool in humans., (© 2024. The Author(s).)

Aims: The study aimed to investigate the role of miR-146a-5p in osteogenesis of hPDLSCs irradiated with low-energy red LEDs., Methods: After irradiation with 5 J/cm 2 red LED, miR-146a-5p expression was detected by real-time quantitative polymerase chain reaction (RT-qPCR), and osteogenic markers expression was determined by RT-qPCR and Western blotting. Alkaline phosphatase (ALP) activity was assessed by ALP staining, and mineralization was assessed by Alizarin Red staining, respectively. Lentiviral vectors were designed to regulate miR-146a-5p expression. Dual-luciferase reporter assay was performed to confirm the targeted relationship between miR-146a-5p and MAPK1. Short hairpin RNA (shRNA) was used to regulate MAPK1 expression., Results: RT-qPCR and western blotting revealed that 5 J/cm 2 irradiation elevated the levels of the osteogenic markers osterix (OSX) and bone sialoprotein (BSP) in hPDLSCs. miR-146a-5p is downregulated in hPDLSCs under the low-energy red LED light irradiation. miR-146a-5p underexpression markedly promoted the osteogenic potential of hPDLSCs. miR-146a-5p targeted MAPK1. 5 J/cm 2 red LED irradiation rescued the inhibitory effects of upregulated miR-146a-5p on osteogenic differentiation, and the positive influence of red LED irradiation could be reversed by downregulated MAPK1., Conclusion: These findings confirm that miR-146a-5p is involved in the effect of LED irradiation on the osteogenic differentiation of hPDLSCs by targeting MAPK1. Red LED irradiation may be a potential clinical adjunct therapy for periodontal regeneration., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Ionizing radiation promotes mammary carcinogenesis. Induction of DNA double-strand breaks (DSBs) is the initial event after radiation exposure, which can potentially lead to carcinogenesis, but the dynamics of DSB induction and repair are not well understood at the tissue level. In this study, we used female rats, which have been recognized as a useful experimental model for studying radiation effects on the mammary gland. We focused on differences in DSB kinetics among basal cells, luminal progenitor and mature cells in different parts of the mammary duct. 53BP1 foci were used as surrogate markers of DSBs, and 53BP1 foci in each mammary epithelial cell in immunostained tissue sections were counted 1-24 h after irradiation and fitted to an exponential function of time. Basal cells were identified as cytokeratin (CK) 14+ cells, luminal progenitor cells as CK8 + 18low cells and luminal mature cells as CK8 + 18high cells. The number of DSBs per nucleus tended to be higher in luminal cells than basal cells at 1 h post-irradiation. A model analysis indicated that basal cells in terminal end buds (TEBs), which constitute the leading edge of the mammary duct, had significantly fewer initial DSBs than the two types of luminal cells, and there was no significant difference in initial amount among the cell types in the subtending duct. The repair rate did not differ among mammary epithelial cell types or their locations. Thus, luminal progenitor and mature cells are more susceptible to radiation-induced DSBs than are basal cells in TEBs., (© The Author(s) 2024. Published by Oxford University Press on behalf of The Japanese Radiation Research Society and Japanese Society for Radiation Oncology.)

Age at exposure is a critical modifier of the risk of radiation-induced cancer. However, the effects of age on radiation-induced carcinogenesis remain poorly understood. In this study, we focused on tissue stem cells using Lgr5-eGFP-ires-Cre ERT2 mice to compare radiation-induced DNA damage responses between Lgr5+ and Lgr5- intestinal stem cells. Three-dimensional immunostaining analyses demonstrated that radiation induced apoptosis and the mitotic index more efficiently in adult Lgr5- stem cells than in adult Lgr5+ stem cells but not in infants, regardless of Lgr5 expression. Supporting this evidence, rapid and transient p53 activation occurred after irradiation in adult intestinal crypts but not in infants. RNA sequencing revealed greater variability in gene expression in adult Lgr5+ stem cells than in infant Lgr5+ stem cells after irradiation. Notably, the cell cycle and DNA repair pathways were more enriched in adult stem cells than in infant stem cells after irradiation. Our findings suggest that radiation-induced DNA damage responses in mouse intestinal crypts differ between infants and adults, potentially contributing to the age-dependent susceptibility to radiation carcinogenesis.

The present study aimed to compare the effect of photobiomodulation with different energy densities on the angiogenic differentiation of human periodontal ligament stem cells (hPDLSCs) and stem cells from human exfoliated deciduous teeth (SHED). Photobiomodulation therapy with a 660 nm diode laser (2.4 J/cm 2 and 3.9 J/cm 2 ) on two consecutive days post-culture was applied to two types of stem cells (hPDLSCs and SHED). The Quantitative Real-time Polymerase Chain Reaction (RT-qPCR) test was undertaken to investigate Vascular Endothelial Growth Factor-A (VEGF-A) and Angiopoietin I (ANG-I) genes on days 1, 3, 5, 7, and 10 after the first session of laser application. The 4',6-diamidino-2-phenylindole (DAPI) staining and Methyl Thiazolyl Tetrazolium (MTT) test were conducted on days 1, 3, and 5 after the first session of laser application, to assess the cell viability. The Two-way ANOVA with Tukey post hoc test was used to analyze the outcomes of the MTT and RT-qPCR tests. The results of the MTT and DAPI convergently illustrated that the groups receiving photobiomodulation with 2.4 J/cm 2 had higher cell viability compared to 3.9 J/cm 2 . All experimental groups showed an upregulation of VEGF-A and ANG-I gene expression from day 1 to 5, followed by a downregulation from day 5 to 10. The groups with cultured hPDLSCs and SHED receiving photobiomodulation using 2.4 J/cm 2 had the most amounts of VEGF-A and ANG-I gene expression on day 5, respectively. In conclusion, the 660 nm mediated photobiomodulation therapy of cultured SHED and hPDLSCs with 2.4 J/cm 2 energy density may be associated with higher angiogenic differentiation (the expression of VEGF-A and ANG-I) as well as higher cell viability compared to the photobiomodulation therapy with 3.9 J/cm 2 ., (© 2024. The Author(s).)

This study aimed to analyze the impact of strontium ranelate (Str), photobiomodulation (PBM), or their combination of the proliferation, osteogenic differentiation, and cementogenic differentiation of buccal fat pad-derived stem cells. BFPdSCs were exposed to one of the following interventions: (1) PBM (660 nm), (2) PBM (660 nm) + Str, (3) PBM (880 nm), (4) PBM (880 nm) + Str, (5) Str. All study groups had significantly higher osteogenic differentiation than the control group (p < 0.05), and no significant difference existed between the 660 and 808 nm groups (p = 0.97). Compared to the Str group, 660 nm and 880 nm group samples had significantly lower osteogenic differentiation (p < 0.0001), while other groups did not show a significant difference. Regarding cementogenic differentiation, the 660 nm group showed higher values than the 808 nm group (p < 0.01). Compared with the Str group, 660 nm, 660 nm + Str, and 808 nm + Str groups showed significantly higher gene expression (p < 0.05). In the case of osteogenic differentiation, although photobiomodulation alone had a lower inducing effect than strontium ranelate, combining 808 nm diode lasers and strontium ranelate may provide the best results. Moreover, using a 660 nm diode laser and exposing stem cells to strontium ranelate can be the most effective approach to induce cementogenic differentiation., (© 2024 American Society for Photobiology.)

Male infertility is a recognized side effect of chemoradiotherapy. Extant spermatogonial stem cells (SSCs) may act as originators for any subsequent recovery. However, which type of SSCs, the mechanism by which they survive and resist toxicity, and how they act to restart spermatogenesis remain largely unknown. Here, we identify a small population of Set domain-containing protein 4 (Setd4)-expressing SSCs that occur in a relatively dormant state in the mouse seminiferous tubule. Extant beyond high-dose chemoradiotherapy, these cells then activate to recover spermatogenesis. Recovery fails when Setd4 + SSCs are deleted. Confirmed to be of fetal origin, these Setd4 + SSCs are shown to facilitate early testicular development and also contribute to steady-state spermatogenesis in adulthood. Upon activation, chromatin remodeling increases their genome-wide accessibility, enabling Notch1 and Aurora activation with corresponding silencing of p21 and p53. Here, Setd4 + SSCs are presented as the originators of both testicular development and spermatogenesis recovery in chemoradiotherapy-induced infertility., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Phototherapy has been extensively used to prevent and treat signs of aging and stimulate wound healing, and phototherapy through light-emitting diodes (LEDs). In contrast to LED, organic LED (OLED) devices are composed of organic semiconductors that possess novel characteristics. We investigated the regenerative potential of OLED for restoring cellular potential from senescence and thus delaying animal aging. Bone marrow-derived stem cells (BMSCs) and adipose-derived stem cells (ADSCs) were isolated from the control and OLED- treated groups to evaluate their proliferation, migration, and differentiation potentials. Cellular senescence was evaluated using a senescence-associated β-galactosidase (SA-β-gal) activity assay and gene expression biomarker assessment. OLED treatment significantly increased the cell proliferation, colony formation, and migration abilities of stem cells. SA-β-gal activity was significantly decreased in both ADSCs and BMSCs in the OLED-treated group. Gene expression biomarkers from treated mice indicated a significant upregulation of IGF-1 (insulin growthfactor-1). The upregulation of the SIRT1 gene inhibited the p16 and p19 genes then to downregulate the p53 expressions for regeneration of stem cells in the OLED-treated group. Our findings indicated that the survival rates of 10-month aging senescence-accelerated mouse prone 8 mice were prolonged and that their gross appearance improved markedly after OLED treatment. Histological analysis of skin and brain tissue also indicated significantly greater collagen fibers density, which prevents ocular abnormalities and β-amyloid accumulation. Lordokyphosis and bone characteristics were observed to resemble those of younger mice after OLED treatment. In conclusion, OLED therapy reduced the signs of aging and enhanced stem-cell senescence recovery and then could be used for tissue regeneration., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: Radiation therapies are often associated with permanent devitalization in the surrounding tissue. The authors hypothesized that stem cells are damaged depending on each irradiation dose and frequency of fractionated radiotherapies, which results in impaired tissue function, including wound-healing capacity., Methods: Susceptibility of human adipose-derived stem cells (ASCs) to a single irradiation (0 to 10 Gy) was assessed in vitro. In vivo chronic radiation effects were also assessed on mouse dorsal skin ( n = 4 to 5) for 6 months after a total of 40 Gy irradiation (0 Gy as control) using 1 of 3 fractionated protocols (2 Gy daily for 20 days, 10 Gy weekly for 4 weeks, or 10 Gy monthly for 4 months). Oxygen partial pressure, oxygen saturation of hemoglobin, and dorsal skin viscoelasticity were measured periodically, and wound healing and tissue immunohistology were compared at 6 months., Results: A single irradiation of cultured human ASCs resulted in a dose-dependent increase in cell death up to 2 Gy but with no further increases between 2 and 10 Gy. Most of the apoptotic ASCs were in the proliferation phase. Among the 3 in vivo irradiation protocols, the 2 Gy×20 group had the most severe chronic tissue damage (ie, skin dysfunction, subcutaneous atrophy, depletion of CD34 + stem cells) 6 months after the irradiation. Wound healing was also impaired most significantly in the 2 Gy×20 group., Conclusion: These results have important clinical implications for surgeons and radiotherapists in the timing of surgical interventions and the optimization of fractionation protocols., Clinical Relevance Statement: Irradiation damages stem cells depending on the radiation dose and frequency. Using the ultimately optimized protocol, surgeons can minimize the long-term functional deficits of radiated tissue without losing the anticancer efficacy of radiation therapy., (Copyright © 2023 by the American Society of Plastic Surgeons.)

Salivary gland homeostasis and regeneration after radiotherapy depend significantly on progenitor cells. However, the lineage of submandibular gland (SMG) progenitor cells remains less defined compared with other normal organs. Here, using a mouse strain expressing regulated CreERT2 recombinase from the endogenous Tert locus, we identify a distinct telomerase-expressing (Tert High ) cell population located in the ductal region of the adult SMG. These Tert High cells contribute to ductal cell generation during SMG homeostasis and to both ductal and acinar cell renewal 1 year after radiotherapy. Tert High cells maintain self-renewal capacity during in vitro culture, exhibit resistance to radiation damage, and demonstrate enhanced proliferative activity after radiation exposure. Similarly, primary human SMG cells with high Tert expression display enhanced cell survival after radiotherapy, and CRISPR-activated Tert in human SMG spheres increases proliferation after radiation. RNA sequencing reveals upregulation of "cell cycling" and "oxidative stress response" pathways in Tert High cells following radiation. Mechanistically, Tert appears to modulate cell survival through ROS levels in SMG spheres following radiation damage. Our findings highlight the significance of Tert High cells in salivary gland biology, providing insights into their response to radiotherapy and into their use as a potential target for enhancing salivary gland regeneration after radiotherapy., (© 2024 Guan et al.; Published by Cold Spring Harbor Laboratory Press.)

Ultraviolet-B (UV-B) radiation overexposure causes function impairment of epidermal stem cells (ESCs). We explored the mechanism of Annexin A1 (ANXA1) ameliorating UV-B-induced ESC mitochondrial dysfunction/cell injury. ESCs were cultured in vitro and irradiated with different doses of UV-B. Cell viability/ANXA1 protein level were assessed. After oe-ANXA1 transfection, ESCs were treated with oe-ANXA1/UV-B irradiation/CCCP/CCG-1423/3-methyladenine for 12 h. Cell viability/death, and adenosine triphosphate (ATP)/reactive oxygen species (ROS) levels were determined. Mitochondrial membrane potential (MMP) changes/DNA (mtDNA) content/oxygen consumption and RhoA activation were assessed. ROCK1/p-MYPT1/MYPT1/(LC3BII/I)/Beclin-1/p62 protein levels were determined. Mitochondrial morphology was observed. Mito-Tracker Green (MTG) and LC3B levels were determined. UV-B irradiation decreased cell viability/ANXA1 expression in a dose-dependent manner. UV-B-treated ESCs exhibited reduced cell viability/ATP content/MMP level/mitochondrial respiratory control ratio/mtDNA number/RhoA activity/MYPT1 phosphorylation/MTG + LC3B + cells/(LC3BII/I) and Beclin-1 proteins, increased cell death/ROS/p62/IL-1β/IL-6/TNF-α expression, contracted mitochondrial, disappeared mitochondrial cristae, and increased vacuolar mitochondria, which were averted by ANXA1 overexpression, suggesting that UV-B induced ESC mitochondrial dysfunction/cell injury/inflammation by repressing mitophagy, but ANXA1 promoted mitophagy by activating the RhoA/ROCK1 pathway, thus repressing UV-B's effects. Mitophagy activation ameliorated UV-B-caused ESC mitochondrial dysfunction/cell injury/inflammation. Mitophagy inhibition partly diminished ANXA1-ameliorated UV-B's effects. Conjointly, ANXA1 promoted mitophagy by activating the RhoA/ROCK1 pathway, thereby improving UV-B-induced ESC mitochondrial dysfunction/cell injury., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Background: Dental pulp stem cells (DPSCs) have self-renewal and multidirectional differentiation potentials. As such, DPSCs have a wide range of clinical applications. Low-level laser therapy (LLLT) has positive photobiostimulatory effects on cell proliferation, angiogenesis, osteogenic differentiation, bone regeneration, and fracture healing. However, there have been few studies on the effect of low-energy lasers on DPSC proliferation., Methods: DPSCs were obtained from dental pulp tissue. The effects of LLLT on the proliferation of DPSCs and the associated mechanisms were investigated by in vitro culture and laser irradiation., Results: LLLT with energy densities of 3.5 J/cm2 and 14 J/cm2promoted the proliferation of DPSCs. Differential protein expression studies suggested the stimulation of DPSC proliferation by LLLT involved the PI3K-Akt and Rap1 signaling pathways, as well as the apoptosis-related pathway., Conclusion: This preliminary study demonstrated that low-energy lasers have a pro-proliferative effect on DPSCs, and identified possible associated mechanisms. Our findings provide a theoretical basis for the clinical application of DPSCs and suggest novel strategies for the treatment of related diseases., Competing Interests: The authors declare no conflict of interest., (© 2024 The Author(s). Published by IMR Press.)

Lysophosphatidic acid (LPA) is a bioactive lipid molecule that regulates a wide array of cellular functions, including proliferation, differentiation, and survival, via activation of cognate receptors. The LPA 5 receptor is highly expressed in the intestinal epithelium, but its function in restoring intestinal epithelial integrity following injury has not been examined. Here, we use a radiation-induced injury model to study the role of LPA 5 in regulating intestinal epithelial regeneration. Control mice ( Lpar5 f/f ) and mice with an inducible, epithelial cell-specific deletion of Lpar5 in the small intestine ( Lpar5 IECKO ) were subjected to 10 Gy total body X-ray irradiation and analyzed during recovery. Repair of the intestinal mucosa was delayed in Lpar5 IECKO mice with reduced epithelial proliferation and increased crypt cell apoptosis. These effects were accompanied by reduced numbers of OLFM4+ intestinal stem cells (ISCs). The effects of LPA 5 on ISCs were corroborated by studies using organoids derived from Lgr5-lineage tracking reporter mice with deletion of Lpar5 in Lgr5+-stem cells (Lgr5 Cont or Lgr5 ΔLpar5 ). Irradiation of organoids resulted in fewer numbers of Lgr5 ΔLpar5 organoids retaining Lgr5+-derived progenitor cells compared with Lgr5 Cont organoids. Finally, we observed that impaired regeneration in Lpar5 IECKO mice was associated with reduced numbers of Paneth cells and decreased expression of Yes-associated protein (YAP), a critical factor for intestinal epithelial repair. Our study highlights a novel role for LPA 5 in regeneration of the intestinal epithelium following irradiation and its effect on the maintenance of Paneth cells that support the stem cell niche. NEW & NOTEWORTHY We used mice lacking expression of the lysophosphatidic acid receptor 5 (LPA 5 ) in intestinal epithelial cells and intestinal organoids to show that the LPA 5 receptor protects intestinal stem cells and progenitors from radiation-induced injury. We show that LPA 5 induces YAP signaling and regulates Paneth cells.

Background: The objective of this study is to investigate the mechanism by which low-level laser stimulation promotes the proliferation of intraepithelial hair follicle stem cells (HFSCs) in wounds. This research aims to expand the applications of laser treatment, enhance wound repair methods, and establish a theoretical and experimental foundation for achieving accelerated wound healing., Methods: The experimental approach involved irradiating a cell model with low-level laser to assess the proliferation of HFSCs and examine alterations in the expression of proteins related to the Wnt/β-catenin signaling pathway. A mouse back wound model was established to investigate the effects of low-level laser irradiation on wound healing rate, wound microenvironment, and the proliferation of HFSCs in relation to the Wnt/β-catenin signaling pathway., Results: The research findings indicate that low-level laser light effectively activates the Wnt signaling pathway, leading to the increased accumulation of core protein β-catenin and the upregulation of key downstream gene Lef 1. Consequently, this regulatory mechanism facilitates various downstream biological effects, including the notable promotion of HFSC proliferation and differentiation into skin appendages and epithelial tissues. As a result, the process of wound healing is significantly accelerated., Conclusion: Low levels of laser activates the Wnt signalling pathway, promotes the regeneration of hair follicle stem cells and accelerates wound healing., (© 2024 The Author(s). Skin Research and Technology published by John Wiley & Sons Ltd.)

Background: Different types of alopecia have negative impacts on patients. Recently, some kinds of laser or light therapies have been reported to effectively alleviate hair loss. Carbon dioxide fractional laser (CO 2 FL) treatment is one of the most effective laser treatments, but its beneficial effects and exact mechanism in hair regrowth have not been reported in detail. The purpose of this study was to investigate the effect and molecular mechanism further., Methods: C57 and Lgr5-Cre: Rosa-mTmG mouse models of hair regrowth were established by CO 2 FL treatment, and the parameters that induced the best effect were determined. Tissues were harvested on the day prior to the treatment day and on days 3, 5, 7, 10 and 14 after CO 2 FL. H&E and immunofluorescence staining, RNA sequencing (RNA-seq), quantitative real-time polymerase chain reaction (qPCR), Western blotting (WB) and related inhibitor were used to determine the molecular mechanism underlying the effect of CO 2 FL treatment on the hair cycle and hair regrowth. In clinical trial, five participants were treated three sessions at 1-month intervals to obverse the effects., Results: Hair regrew and covered the treatment area on the tenth day after CO 2 FL treatment with the best parameters, while the control group showed signs of hair growth on the 14th day. H&E and immunofluorescence staining showed that the transition of hair follicles (HFs) from telogen to anagen was accelerated, and the rapid activation and proliferation of Lgr5+ hair follicle stem cells (HFSCs) were observed in the treatment group. The RNA-seq, qPCR and WB results indicated that the Wnt pathway was significantly activated after CO 2 FL treatment. Improvement achieved with CO 2 FL treatment in clinical trial., Conclusions: The results of this study suggest that CO 2 FL treatment can promote hair regrowth by activating Lgr5+ HFSCs and upregulating the Wnt/β-catenin pathway. Clinical trial results demonstrated that CO 2 FL treatment will be a promising therapeutic regimen for alopecia., No Level Assigned: This journal requires that authors assign a level of evidence to each submission to which Evidence-Based Medicine rankings are applicable. This excludes Review Articles, Book Reviews, and manuscripts that concern Basic Science, Animal Studies, Cadaver Studies, and Experimental Studies. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 ., (© 2023. Springer Science+Business Media, LLC, part of Springer Nature and International Society of Aesthetic Plastic Surgery.)

Photobiomodulation therapy (PBMT) is the application of a low-level laser device to generate physiological changes and provide therapeutic effects. Till now, the effects of PBMT on the neural differentiation of mesenchymal stem cells have been rarely reported. Herein, the potential effect and mechanism of PBMT on the neural differentiation of dental pulp stem cells (DPSCs) were preliminarily investigated in our research. The optimal dose of 3.75 J/cm 2 was first screened for use in the following neural-inducing studies. Then, DPSCs were cultured in neural induction medium and treated with laser irradiation for 7 days. From the results of morphology and immunofluorescence, we found that irradiation promoted the formation of neural stem cell-like spheroids derived from DPSCs and enhanced potential neural differentiation. Furthermore, neural differentiation gene expressions of Nestin, microtubule-associated protein-2, and neural cell adhesion molecule were increased after PBMT irradiation. The protein expressions of class III β-tubulin and neurogenic differentiation factor 1 were also improved. Meanwhile, the involvement of extracellular signal-regulated kinase (ERK1/2) was investigated by western blot. Our study showed that the neural differentiation of DPSCs was promoted by PBMT, and the underlying mechanism in this process was associated with activating the ERK1/2 signaling pathway., (© 2023 American Society for Photobiology.)

One of the biggest challenges in tissue engineering and regenerative medicine is to ensure oxygen supply of cells in the (temporary) absence of vasculature. With the vision to exploit photosynthetic oxygen production by microalgae, co-cultivated in close vicinity to oxygen-consuming mammalian cells, we are searching for culture conditions that are compatible for both sides. Herein, we investigated the impact of long-term illumination on mammalian cells which is essential to enable photosynthesis by microalgae: four different cell types-primary human fibroblasts, dental pulp stem cells, and osteoblasts as well as the murine beta-cell line INS-1-were continuously exposed to warm white light, red or blue light over seven days. We observed that illumination with red light has no adverse effects on viability, metabolic activity and growth of the cells whereas exposure to white light has deleterious effects that can be attributed to its blue light portion. Quantification of intracellular glutathione did not reveal a clear correlation of this effect with an enhanced production of reactive oxygen species. Finally, our data indicate that the cytotoxic effect of short-wavelength light is predominantly a direct effect of cell illumination; photo-induced changes in the cell culture media play only a minor role., (© 2024. The Author(s).)

Background: This investigation set out to compare the impacts of low-level diode laser (LLDL) and red light-emitting diode (LED) on the survival of human dental pulp stem cells (hDPSCs) and osteogenic/odontogenic differentiation. Methods and materials: In this ex vivo experimental study, the experimental groups underwent the irradiation of LLDL (4 J/cm 2 energy density) and red LED in the osteogenic medium. Survival of hDPSCs was assessed after 24 and 48 h ( n  = 9) using the methyl thiazolyl tetrazolium (MTT) assay. The assessment of osteogenic/odontogenic differentiation was conducted using alizarin red staining (ARS; three repetitions). The investigation of osteogenic and odontogenic gene expression was performed at two time points, specifically 24 and 48 h ( n  = 12). This analysis was performed utilizing real-time reverse-transcription polymerase chain reaction (RT-PCR). The groups were compared at each time point using SPSS version 24. To analyze the data, the Mann-Whitney U test, analysis of variance, Tukey's test, and t -test were utilized. Results: The MTT assay showed that LLDL significantly decreased the survival of hDPSCs after 48 h, compared with other groups ( p  < 0.05). The qualitative results of ARS revealed that LLDL and red LED increased the osteogenic differentiation of hDPSCs. LLDL and red LED both upregulated the expression of osteogenic/odontogenic genes, including bone sialoprotein (BSP), alkaline phosphatase (ALP), dentin matrix protein 1 (DMP1), and dentin sialophosphoprotein (DSPP), in hDPSCs. The LLDL group exhibited a higher level of gene upregulation ( p  < 0.0001). Conclusions: The cell survival of hDPSCs was reduced, despite an increase in osteogenic/odontogenic activity. Clinical relevance: Introduction of noninvasive methods in regenerative endodontic treatments.

Background and Purpose: Patients undergoing radiotherapy for head and neck cancer often experience a decline in their quality of life due to the co-irradiation of salivary glands. Radiation-induced cellular senescence is a key factor contributing to salivary gland dysfunction. Interestingly, mitochondrial dysfunction and cellular senescence have been reported to be strongly interconnected and thus implicated in several aging-related diseases. This study aims to investigate the role of mitochondrial dysfunction in senescence induction in salivary gland stem/progenitor cells after irradiation., Materials and Methods: A dose of 7 Gy photons was used to irradiate mouse salivary gland organoids. Senescent markers and mitochondrial function were assessed using rt-qPCR, western blot analysis, SA-β-Gal staining and flow cytometry analysis. Mitochondrial dynamics-related proteins were detected by western blot analysis while Mdivi-1 and MFI8 were used to modulate the mitochondrial fission process. To induce mitophagy, organoids were treated with Urolithin A and PMI and subsequently stem/progenitor cell self-renewal capacity was assessed as organoid forming efficiency., Results: Irradiation led to increased senescence and accumulation of dysfunctional mitochondria. This was accompanied by a strong downregulation of mitochondrial fission-related proteins and mitophagy-related genes. After irradiation, treatment with the mitophagy inducer Urolithin A attenuated the senescent phenotype and improved organoid growth and stem/progenitor cell self-renewal capacity., Conclusion: This study shows the important interplay between senescence and mitochondrial dysfunction after irradiation. Importantly, activation of mitophagy improved salivary gland stem/progenitor cell function thereby providing a novel therapeutic strategy to restore the regenerative capacity of salivary glands following irradiation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

Purpose: Increasing epidemiological and biological evidence suggests that radiation exposure enhances cancer risk in a dose-dependent manner. This can be attributed to the 'dose-rate effect,' where the biological effect of low dose-rate radiation is lower than that of the same dose at a high dose-rate. This effect has been reported in epidemiological studies and experimental biology, although the underlying biological mechanisms are not completely understood. In this review, we aim to propose a suitable model for radiation carcinogenesis based on the dose-rate effect in tissue stem cells., Methods: We surveyed and summarized the latest studies on the mechanisms of carcinogenesis. Next, we summarized the radiosensitivity of intestinal stem cells and the role of dose-rate in the modulation of stem-cell dynamics after irradiation., Results: Consistently, driver mutations can be detected in most cancers from past to present, supporting the hypothesis that cancer progression is initiated by the accumulation of driver mutations. Recent reports demonstrated that driver mutations can be observed even in normal tissues, which suggests that the accumulation of mutations is a necessary condition for cancer progression. In addition, driver mutations in tissue stem cells can cause tumors, whereas they are not sufficient when they occur in non-stem cells. For non-stem cells, tissue remodeling induced by marked inflammation after the loss of tissue cells is important in addition to the accumulation of mutations. Therefore, the mechanism of carcinogenesis differs according to the cell type and magnitude of stress. In addition, our results indicated that non-irradiated stem cells tend to be eliminated from three-dimensional cultures of intestinal stem cells (organoids) composed of irradiated and non-irradiated stem cells, supporting the stem-cell competition., Conclusions: We propose a unique scheme in which the dose-rate dependent response of intestinal stem cells incorporates the concept of the threshold of stem-cell competition and context-dependent target shift from stem cells to whole tissue. The concept highlights four key issues that should be considered in radiation carcinogenesis: i.e. accumulation of mutations; tissue reconstitution; stem-cell competition; and environmental factors like epigenetic modifications.

The effect of near infrared (NIR) laser irradiation on proliferation and osteogenic differentiation of buccal fat pad-derived stem cells and the role of transient receptor potential (TRP) channels was investigated in the current research. After stem cell isolation, a 940 nm laser with 0.1 W, 3 J/cm 2 was used in pulsed and continuous mode for irradiation in 3 sessions once every 48 h. The cells were cultured in the following groups: non-osteogenic differentiation medium/primary medium (PM) and osteogenic medium (OM) groups with laser-irradiated (L +), without irradiation (L -), laser treated + Capsazepine inhibitor (L + Cap), and laser treated + Skf96365 inhibitor (L + Skf). Alizarin Red staining and RT-PCR were used to assess osteogenic differentiation and evaluate RUNX2, Osterix, and ALP gene expression levels. The pulsed setting showed the best viability results (P < 0.05) and was used for osteogenic differentiation evaluations. The results of Alizarin red staining were not statistically different between the four groups. Osterix and ALP expression increased in the (L +) group. This upregulation abrogated in the presence of Capsazepine, TRPV1 inhibitor (L + Cap); however, no significant effect was observed with Skf96365 (L + Skf)., (© 2022. The Author(s), under exclusive licence to Springer-Verlag London Ltd., part of Springer Nature.)

Radiation response differs depending on the dose and dose rate in intestinal stem cells; however, the underlying mechanisms are not clear. To understand the effects of low-dose and low-dose-rate radiation, the authors established an organoid system that mimics the in vivo environment and sporadic low-dose-rate irradiation conditions in vitro. Organoid-forming potential and the number of stem cells in the organoids derived from 1 Gy-irradiated cells were lower than those from non-irradiated cells; however, the difference was not significant, although 1 Gy-irradiated stem cells exhibited significant growth disadvantage in the mixed-organoid with non-irradiated and irradiated stem cells. Furthermore, the authors irradiated a cell with X-ray microbeams and performed time-lapse observations and found that irradiated cells did not remain in the organoid. These results suggest that radiation-induced stem cell competition can occur in intestinal organoids and contribute to a low risk of cancers at low-dose-rate exposures., (© The Author(s) 2022. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

The uncertain cancer risk of protracted radiation exposure at low dose rates is an important issue in radiological protection. Tissue stem/progenitor cells are a supposed origin of cancer and may contribute to the dose-rate effect on carcinogenesis. The authors have shown that female rats subjected to continuous whole body γ irradiation as juveniles or young adults have a notably reduced incidence of mammary cancer as compared with those irradiated acutely. Experiments using the mammosphere formation assay suggested the presence of radioresistant progenitor cells. Cell sorting indicated that basal progenitor cells in rat mammary gland were more resistant than luminal progenitors to killing by acute radiation, especially at high doses. Thus, the evidence indicates a cell-type-dependent inactivation of mammary cells that manifests only at high acute doses, implying a link to the observed dose-rate effect on carcinogenesis., (© The Author(s) 2022. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Objective: In recent years, fractionated irradiation protocols, rather than a simple plan of exposure, have been proposed as a more effective method in the field of tissue regeneration. Thus, this study aimed at a comparative analysis of single versus double irradiation of an 808-nm diode laser, in terms of dental pulp stem cells' (DPSCs) viability and proliferation in vitro . Methods: Subcultured DPSCs were either irradiated, or not (control group), with energy densities of 3, 7, and 12 J·cm -2 in a single- or double-session manner (24 h apart). On 0, 12, 24, 48, and 72 h postirradiation, cell viability and proliferation were evaluated through Trypan Blue and alamarBlue assays, respectively. Results: During the first 48 h postirradiation, the highest rates of DPSC proliferation were assigned to double irradiation at 3 or single exposure to 7 J⋅cm -2 , with no cytotoxic effects on cell viability. Inversely, single irradiation at 12, or a double session of exposure to 7 or 12 J⋅cm -2 , led to a significant descent in the rates of proliferation and cell viability. Conclusions: Within the limitations of this study, evidence suggests a positive impact on the biological responses of DPSCs following double session of exposure to lower energy densities as well as a single irradiation at a higher energy dosage.

Intestinal stem cells (ISCs) are essential for the regeneration of intestinal cells upon radiation or chemical agent damage. As for radiation-induced damage, the expression of AIM2, YAP, TLR3, PUMA or BVES can aggravate ISCs depletion, while the stimulation of TLR5, HGF/MET signaling, Ass1 gene, Slit/Robo signaling facilitate the radio-resistance of ISCs. Upon chemical agent treatment, the activation of TRAIL or p53/PUMA pathway exacerbate injury on ISCs, while the increased levels of IL-22, β-arrestin1 can ease the damage. The transformation between reserve ISCs (rISCs) maintaining quiescent states and active ISCs (aISCs) that are highly proliferative has obtained much attention in recent years, in which ISCs expressing high levels of Hopx, Bmi1, mTert, Krt19 or Lrig1 are resistant to radiation injury, and SOX9, MSI2, clusterin, URI are essential for rISCs maintenance. The differentiated cells like Paneth cells and enteroendocrine cells can also obtain stemness driven by radiation injury mediated by Wnt or Notch signaling. Besides, Mex3a-expressed ISCs can survive and then proliferate into intestinal epithelial cells upon chemical agent damage. In addition, the modulation of symbiotic microbes harboring gastrointestinal (GI) tract is also a promising strategy to protect ISCs against radiation damage. Overall, the strategies targeting mechanisms modulating ISCs activities are conducive to alleviating GI injury of patients receiving chemoradiotherapy or victims of nuclear or chemical accident., (© The Author(s) 2022. Published by Oxford University Press on behalf of The Japanese Radiation Research Society and Japanese Society for Radiation Oncology.)

Intestinal organoids are an in vitro cultured tissue model generated from intestinal stem cells, and they contain a mixture of epithelial cell types. We previously established an efficient 'one cell/well' sorting method, and defined organoid-forming potential (OFP) as a useful index to evaluate the stemness of individual cells. In this study, we assessed the response to radiation dose and dose-rate by measuring both OFP and the percentage of stem cells in the crypts. After high-dose-rate (HDR, 0.5 Gy/min) irradiation in vivo, the percentage of stem cells in the harvested crypt cells decreased, and the replenishment of cycling stem cells originating from dormant cells was enhanced, but OFP increased in cells irradiated with a total dose of >1 Gy. In contrast, at a total dose of 0.1 Gy the percentage of stem cells reduced slightly, but neither replenishment rate nor OFP changed. Furthermore, the response to 1 Gy of low-dose-rate (LDR) irradiation was similar to the response to 0.1 Gy HDR irradiation. These results suggest that 0.1 Gy HDR irradiation or 1 Gy LDR irradiation does not alter stemness. Additionally, the OFP increase in the colon in response to irradiation was smaller than that in the duodenum, similar to the percentage of stem cells. Understanding the differences in the response of stem cells between the colon and the duodenum to radiation is important to clarify the mechanisms underlying the development of radiation-associated intestinal cancers., (© The Author(s) 2021. Published by Oxford University Press on behalf of The Japanese Radiation Research Society and Japanese Society for Radiation Oncology.)

Objective: To determine the effect of different energy densities of near infrared diode lasers with wavelengths of 810 or 940 nm on the proliferation and survival of periodontal ligament derived stem cells (PDLSCs)., Methods: After isolation and characterisation, PDLSCs were cultured in clear 96-well plates. Each well was irradiated by either 810 nm (L1) or 940 nm (L2) lasers, with energy densities of 0.5, 1.5 and 2.5 J/cm2 and an output power of 100 mW. A non-irradiated well was used as a control. Cellular viability was measured 24 hours after irradiation using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay and proliferation was measured 24, 48 and 72 hours after irradiation using trypan blue staining and counting. Propidium iodide (PI) staining was used to identify any pyknotic nuclei or nuclear fragmentation 72 hours after irradiation., Results: An increase in viability was observed only in the group with the 940 nm laser irradiation at energy density of 2.5 J/cm2 (P < 0.001). The proliferation of cells was significantly increased in the group with 940 nm laser irradiation at energy density of 2.5 J/cm2 at all the time points examined in comparison to other groups (P < 0.001). PI staining showed no change in cell nuclei in any of the groups., Conclusion: Irradiation of PDLSCs with a 940 nm laser at an energy density of 2.5 J/cm2 could promote efficient cell proliferation.

The influence of high-linear energy transfer (LET) particle radiation on the functionalities of mesenchymal stromal cells (MSCs) is largely unknown. Here, we analyzed the effects of proton ( 1 H), helium ( 4 He), carbon ( 12 C) and oxygen ( 16 O) ions on human bone marrow-MSCs. Cell cycle distribution and apoptosis induction were examined by flow cytometry, and DNA damage was quantified using γH2AX immunofluorescence and Western blots. Relative biological effectiveness values of MSCs amounted to 1.0-1.1 for 1 H, 1.7-2.3 for 4 He, 2.9-3.4 for 12 C and 2.6-3.3 for 16 O. Particle radiation did not alter the MSCs' characteristic surface marker pattern, and MSCs maintained their multi-lineage differentiation capabilities. Apoptosis rates ranged low for all radiation modalities. At 24 h after irradiation, particle radiation-induced ATM and CHK2 phosphorylation as well as γH2AX foci numbers returned to baseline levels. The resistance of human MSCs to high-LET irradiation suggests that MSCs remain functional after exposure to moderate doses of particle radiation as seen in normal tissues after particle radiotherapy or during manned space flights. In the future, in vivo models focusing on long-term consequences of particle irradiation on the bone marrow niche and MSCs are needed., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Exogenous electrical fields have been explored in regenerative medicine to increase cellular expression of pro-regenerative growth factors. Adipose-derived stem cells (ASCs) are attractive for regenerative applications, specifically for neural repair. Little is known about the relationship between low-level electrical stimulation (ES) and ASC regenerative potentiation. In this work, patterns of ASC expression and secretion of growth factors (i.e., secretome) were explored across a range of ES parameters. ASCs were stimulated with low-level stimulation (20 mV/mm) at varied pulse frequencies, durations, and with alternating versus direct current. Frequency and duration had the most significant effects on growth factor expression. While a range of stimulation frequencies (1, 20, 1000 Hz) applied intermittently (1 h × 3 days) induced upregulation of general wound healing factors, neural-specific factors were only increased at 1 Hz. Moreover, the most optimal expression of neural growth factors was achieved when ASCs were exposed to 1 Hz pulses continuously for 24 h. In evaluation of secretome, apparent inconsistencies were observed across biological replications. Nonetheless, ASC secretome (from 1 Hz, 24 h ES) caused significant increase in neurite extension compared to non-stimulated control. Overall, ASCs are sensitive to ES parameters at low field strengths, notably pulse frequency and stimulation duration., (© 2021. Biomedical Engineering Society.)

Low-intensity pulsed ultrasound (LIPUS) is a promising therapy that has been increasingly explored in basic research and clinical applications. LIPUS is an appealing therapeutic option as it is a noninvasive treatment that has many advantages, including no risk of infection or tissue damage and no known adverse reactions. LIPUS has been shown to have many benefits including promotion of tissue healing, angiogenesis, and tissue regeneration; inhibition of inflammation and pain relief; and stimulation of cell proliferation and differentiation. The biophysical mechanisms of LIPUS remain unclear and the studies are ongoing. In recent years, more and more research has focused on the relationship between LIPUS and stem/progenitor cells. A comprehensive search of the PubMed and Embase databases to July 2020 was performed. LIPUS has many effects on stem cells. Studies show that LIPUS can stimulate stem cells in vitro; promote stem cell proliferation, differentiation, and migration; maintain stem cell activity; alleviate the problems of insufficient seed cell source, differentiation, and maturation; and circumvent the low efficiency of stem cell transplantation. The mechanisms involved in the effects of LIPUS are not fully understood, but the effects demonstrated in studies thus far have been favorable. Much additional research is needed before LIPUS can progress from basic science research to large-scale clinical dissemination and application., Competing Interests: None

Small extracellular vesicles (sEV) facilitate signaling molecule transfer among cells. We examined the therapeutic efficacy of human dental pulp stem cell-derived sEV (hDPSC-sEV) against cellular senescence in an irradiated-submandibular gland mouse model. Seven-week-old mice were exposed to 25 Gy radiation and randomly assigned to control, phosphate-buffered saline (PBS), or hDPSC-sEV groups. At 18 days post-irradiation, saliva production was measured; histological and reverse transcription-quantitative PCR analyses of the submandibular glands were performed. The salivary flow rate did not differ significantly between the PBS and hDPSC-sEV groups. AQP5-expressing acinar cell numbers and AQP5 expression levels in the submandibular glands were higher in the hDPSC-sEV group than in the other groups. Furthermore, compared with non-irradiated mice, mice in the 25 Gy + PBS group showed a high senescence-associated-β-galactosidase-positive cell number and upregulated senescence-related gene (p16 INK4a , p19 Arf , p21) and senescence-associated secretory phenotypic factor (MMP3, IL-6, PAI-1, NF-κB, and TGF-β) expression, all of which were downregulated in the hDPSC-sEV group. Superoxide dismutase levels were lower in the PBS group than in the hDPSC-sEV group. In summary, hDPSC-sEV reduced inflammatory cytokine and senescence-related gene expression and reversed oxidative stress in submandibular cells, thereby preventing irradiation-induced cellular senescence. Based on these results, we hope to contribute to the development of innovative treatment methods for salivary gland dysfunction that develops after radiotherapy for head and neck cancer., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Radiotherapy (RT) is one of the cornerstones in the current treatment paradigm for glioblastoma (GBM). However, little has changed in the management of GBM since the establishment of the current protocol in 2005, and the prognosis remains grim. Radioresistance is one of the hallmarks for treatment failure, and different therapeutic strategies are aimed at overcoming it. Among these strategies, nanomedicine has advantages over conventional tumor therapeutics, including improvements in drug delivery and enhanced antitumor properties. Radiosensitizing strategies using nanoparticles (NP) are actively under study and hold promise to improve the treatment response. We aim to describe the basis of nanomedicine for GBM treatment, current evidence in radiosensitization efforts using nanoparticles, and novel strategies, such as preoperative radiation, that could be synergized with nanoradiosensitizers.

Human skin is a highly efficient self-renewing barrier that is critical to withstanding environmental insults. Undifferentiated keratinocyte stem cells reside in the basal layer of the epidermis and in hair follicles that continuously give rise to progenies ensuring epidermal turnover and renewal. Ultraviolet (UV) radiation is a proven cause of skin keratinocyte cancers, which preferentially occur at sun-exposed areas of the skin. Fortunately, melanocytes produce melanin that is packaged in specific organelles (termed melanosomes) that are then delivered to nearby keratinocytes, endowing the recipient cells with photoprotection. It has long been thought that melanosome transfer takes place stochastically from melanocytes to keratinocytes via an as-yet-unrecognized manner. However, recent studies have indicated that melanosomes are distributed regionally in the basal layer of the skin, affording localized intensive photoprotection for progenitor keratinocytes and stem cells that reside in the microenvironment of the basal epidermis. In this review, we summarize current knowledge about molecular and cellular mechanisms that are responsible for the selective transfer and exclusive degradation of melanosomes in the epidermis, emphasizing implications for skin carcinogenesis., (Copyright © 2021 Japanese Society for Investigative Dermatology. Published by Elsevier B.V. All rights reserved.)

In this study, we evaluated cardiomyogenic differentiation of electromechanically stimulated rat bone marrow-derived stem cells (rt-BMSCs) on an acellular bovine pericardium (aBP) and we looked at the functioning of this engineered patch in a rat myocardial infarct (MI) model. aBP was prepared using a detergent-based decellularization procedure followed by rt-BMSCs seeding, and electrical, mechanical, or electromechanical stimulations (3 millisecond pulses of 5 V cm -1 at 1 Hz, 5% stretching) to enhance cardiomyogenic differentiation. Furthermore, the electromechanically stimulated patch was applied to the MI region over 3 weeks. After this period, the retrieved patch and infarct region were evaluated for the presence of calcification, inflammatory reaction (CD68), patch to host tissue cell migration, and structural sarcomere protein expressions. In conjunction with any sign of calcification, a higher number of BrdU-labelled cells, and a low level of CD68 positive cells were observed in the infarct region under electromechanically stimulated conditions compared with static conditions. More importantly, MHC, SAC, Troponin T, and N-cad positive cells were observed in both infarct region, and retrieved engineered patch after 3 weeks. In a clear alignment with other results, our developed acellular patch promoted the expression of cardiomyogenic differentiation factors under electromechanical stimulation. Our engineered patch showed a successful integration with the host tissue followed by the cell migration to the infarct region., (© 2021 IOP Publishing Ltd.)

Purpose: Radiation-induced gastrointestinal syndrome (RIGS) is currently the main cause of death for people exposed to a high dose of irradiation during nuclear incidents, and there is currently no approved effective therapy. Here, we found that CBP/P300 inhibitors, with high efficacy and low toxicity, might be promising radiation mitigators that can cure RIGS., Methods and Materials: Ex vivo 3D organoid cultures derived from mouse jejunum and human ileum and colon were used to examine the radio-mitigative effects of CBP/P300 inhibitors. The radio-mitigative effect was evaluated by quantifying the survival rate and size of organoids after radiation. SGC-CBP30 (50 mg/kg body weight), an inhibitor of CBP/P300, was intraperitoneally injected into C57B/6J mice 24 hours after subtotal-body irradiation or whole-body irradiation. The regenerated crypts and animal survival were determined by microcolony assay and the Kaplan-Meier method, respectively. Lgr5-lacZ mice were used to evaluate the survival of intestinal stem cells after treatments., Results: We found that CBP/P300 inhibitors were effective mitigators that could be used to treat RIGS. CBP/P300 inhibition promoted the regeneration of intestinal organoids in vitro and of crypts in vivo. Remarkably, the administration of CBP/P300 inhibitors to mice 24 hours after lethal irradiation promoted Lgr5 + intestinal stem cell and crypt recovery, resulting in improved mouse survival. Moreover, our data show that CBP/P300 inhibitors rescued irradiated mice from RIGS by delaying intestinal epithelial cell cycle progression after radiation., Conclusions: These data demonstrate that CBP/P300 inhibitors are effective medical countermeasures to mitigate gastrointestinal toxicity from radiation., (Copyright © 2021 Elsevier Inc. All rights reserved.)

In this article, we provide an extensive review of the recent literature of the signaling pathways modulated by Pulsed Electromagnetic Fields (PEMFs) and PEMFs clinical application. A review of the literature was performed on two medical electronic databases (PubMed and Embase) from 3 to 5 March 2021. Three authors performed the evaluation of the studies and the data extraction. All studies for this review were selected following these inclusion criteria: studies written in English, studies available in full text and studies published in peer-reviewed journal. Molecular biology, identifying cell membrane receptors and pathways involved in bone healing, and studying PEMFs target of action are giving a solid basis for clinical applications of PEMFs. However, further biology studies and clinical trials with clear and standardized parameters (intensity, frequency, dose, duration, type of coil) are required to clarify the precise dose-response relationship and to understand the real applications in clinical practice of PEMFs.

We used light to irradiate skin-derived stem cells and tried to find any cellular protein alterations 24 h after illumination. A 266-nm laser with four intensities was used, and of the nine cell markers that were surveyed in our trials, only CD90 was downregulated at an intensity of 20 μJ for 10 s. Repeated illuminations from the 266-nm laser at seven intensities revealed that CD90 expression was downregulated 14.6-28.8%, depending on light intensity. The maximal effect was noted at an intensity of 30 μJ for 2 s. This innovative finding reveals that a 266-nm laser can regulate protein expression in skin-derivative stem cells.

Human periodontal ligament stem cells (hPDLSCs) sheets play an important role in periodontal tissue engineering. Low-intensity pulsed ultrasound (LIPUS) has been reported as an effective stimulus to regulate cell biological behavior. The present study aims to explore the potential of LIPUS to promote the formation and function of hPDLSC sheets (hPDLSCSs). Hematoxylin-eosin (H&E) staining, western blot, real-time PCR, alkaline phosphatase (ALP), and alizarin red staining were used to evaluate the formation and osteogenic effect of LIPUS on hPDLSCSs in vitro. Hydroxyapatite with or without hPDLSCSs was transplanted in the subcutaneous pockets on the back of nude mice and histological analysis was performed. H&E staining showed increased synthesis of extracellular matrix (ECM) and real-time PCR detected a significant increase in ECM-related genes after LIPUS treatment. In addition, LIPUS could promote the expression of osteogenic differentiation-related genes and proteins. ALP and alizarin red staining also found LIPUS enhanced the osteogenesis of hPDLSCSs. After transplantation in vivo, more dense collagen fibers similar to periodontal ligament were regenerated. Collectively, these results indicate that LIPUS not only promotes the formation and osteogenic differentiation of hPDLSCSs but also is a potential treatment strategy for periodontal tissue engineering., (© 2020 Wiley Periodicals LLC.)

Ultraviolet B (UVB) radiation induces mutagenic DNA photolesions in skin cells especially in form of cyclobutane pyrimidine dimers (CPDs). Protection mechanisms as DNA repair and apoptosis are of great importance in order to prevent skin carcinogenesis. In human skin, neural crest-derived precursors of melanocytes, the dermal stem cells (DSCs), are discussed to be at the origin of melanoma. Although they are constantly exposed to solar UV radiation, it is still not investigated how DSCs cope with UV-induced DNA damage. Here, we report a comparative study of the DNA damage response after irradiation with a physiological relevant UVB dose in DSCs in comparison to fibroblasts, melanocytes and keratinocytes isolated from human foreskin. Within our experimental settings, DSCs were able to repair DNA photolesions as efficient as the other skin cell types with solely keratinocytes repairing significantly faster. Interestingly, only fibroblasts showed significant alterations in cell cycle distribution in terms of a transient S phase arrest following irradiation. Moreover, with the applied UVB dose none of the examined cell types was prone to UVB-induced apoptosis. This may cause persistent genomic alterations and in case of DSCs it may have severe consequences for their daughter cells, the differentiated melanocytes. Altogether, this is the first study demonstrating a similar UV response in dermal stem cells compared to differentiated skin cells., (Copyright © 2021 Elsevier B.V. All rights reserved.)

It is unclear why medulloblastoma patients receiving similar treatments experience different outcomes. Transcriptomic profiling identified subgroups with different prognoses, but in each subgroup, individuals remain at risk of incurable recurrence. To investigate why similar-appearing tumors produce variable outcomes, we analyzed medulloblastomas triggered in transgenic mice by a common driver mutation expressed at different points in brain development. We genetically engineered mice to express oncogenic SmoM2, starting in multipotent glio-neuronal stem cells, or committed neural progenitors. Both groups developed medulloblastomas with similar transcriptomic profiles. We compared medulloblastoma progression, radiosensitivity, and cellular heterogeneity, determined by single-cell transcriptomic analysis (scRNA-seq). Stem cell-triggered medulloblastomas progressed faster, contained more OLIG2-expressing stem-like cells, and consistently showed radioresistance. In contrast, progenitor-triggered MBs progressed slower, down-regulated stem-like cells and were curable with radiation. Progenitor-triggered medulloblastomas also contained more diverse stromal populations, with more Ccr2+ macrophages and fewer Igf1+ microglia, indicating that developmental events affected the subsequent tumor microenvironment. Reduced mTORC1 activity in M-Smo tumors suggests that differential Igf1 contributed to differences in phenotype. Developmental events in tumorigenesis that were obscure in transcriptomic profiles thus remained cryptic determinants of tumor composition and outcome. Precise understanding of medulloblastoma pathogenesis and prognosis requires supplementing transcriptomic/methylomic studies with analyses that resolve cellular heterogeneity.