• ADMSC
• DNA repair
• NHEJ
• RNAseq
• Stem cells

• Cell migration
• DNA damage
• Mesenchymal stem cells
• Osteoblasts
• Radiation backscatter
• Titanium implants

• Humans
• Dental Implants
• Titanium/pharmacology
• Radiation, Ionizing
• Cell Movement
• Plastics

• 6902696 Kreftforeningen
• I-AS-15-050 Dentsply Sirona
• University of Oslo (incl Oslo University Hospital)

• canine
• clinical safety
• exosome
• hemocompatibility
• human
• mesenchymal stromal cells
• tissue factor

• na Terry Johnson Center for Basic Cancer Research
• NA State of Kansas

• immunology profile
• ionizing radiation
• low dose
• mesenchymal stromal cells
• proliferation
• secretory profile

• Adult
• Humans
• Mesenchymal Stem Cells/physiology
• Radiation, Ionizing
• Mercury
• Russia
• Cells, Cultured
• Cell Differentiation

• immuomodulation
• inflammatory diseases
• mesenchymal stem cells
• pretreatment
• secretome

It has been known that senescence-associated secretory phenotype (SASP) triggers senescence of the surrounding normal cells. However, SASP signaling regarding mesenchymal stromal cell aging remains to be fully elucidated. Therefore, the present study aimed to clarify the molecular mechanism of late (passage) MSC-induced paracrine SASP-mediated senescence of early (passage) MSCs during ex vivo expansion. Here, we conducted an extensive characterization of senescence features in bone-marrow (BM)-derived MSCs from healthy human donors. Late MSCs displayed an enlarged senescent-like morphology, induced SASP-related proinflammatory cytokines (IL-1α and IL-8), and reduced clonogenic capacity and osteogenic differentiation when compared to early MSCs. Of note, paracrine effects of SASP-related IL-1α and IL-8 from late MSCs induced cellular senescence of early MSCs via an NF-κB-dependent manner. Moreover, cellular senescence of early MSCs was promoted by the synergistic action of IL-1α and IL-8. However, inhibition of NF-κB by shRNA transfection or using inhibitors in early MSCs blocked early MSCs cellular senescence caused by paracrine SASP of late MSCs. In conclusion, these findings reveal that late MSCs display features of senescence and that, during ex vivo expansion, SASP-related proinflammatory cytokines contribute to activate a cellular senescence program in early MSCs that may ultimately impair their functionality.

• IL-1α
• IL-8
• NF-κB
• mesenchymal stromal cells
• paracrine
• senescence
• senescence-associated secretory phenotype (SASP)

• clinical trials
• extracellular vesicles (EVs)
• mesenchymal stem cells
• regenerative therapy
• single cell transcriptomics
• umbilical cord (UC)

• R01 DK119293 NIDDK NIH HHS
• National Institute of Diabetes and Digestive and Kidney Diseases
• Stanford Maternal and Child Health Research Institute

• bone sarcoma
• cartilage cryoinjury
• cartilage cryopreservation
• osteoarticular reconstruction
• vitrification

• Biomarkers
• Cell Differentiation
• Cellular Senescence/physiology
• Mesenchymal Stem Cells

• National Natural Science Foundation of China
• Sichuan University

• HaCaT
• RNA-seq
• arsenic
• passage matching
• pathway analysis

Alopecia areata (AA) is a common autoimmune disease characterized by hair loss. AA appears in extensive forms, such as progressive and diffusing hair loss (diffuse AA), a total loss of scalp hair (alopecia totalis), and complete loss of hair over the entire body (alopecia universalis). Recently, mesenchymal stem cells (MSCs) have been identified as a therapeutic alternative for autoimmune diseases. For this reason, preclinical and case studies of AA and related diseases using MSCs have been conducted.

Case 1: A 55-year-old woman suffered from AA in two areas of the scalp. She was given 15 rounds of minimally manipulated umbilical cord-MSCs (MM-UC-MSCs) over 6 mo. The AA gradually improved 3 mo after the first round. The patient was cured, and AA did not recur. Case 2: A 30-year-old woman, with history of local steroid hormone injections, suffered from AA in one area on the scalp. She was given two rounds of MM-UC-MSCs over 1 mo. The AA immediately improved after the first round. The patient was cured, and AA did not recur. Case 3: A 20-year-old woman, who was diagnosed with alopecia universalis at the age of 12, was given 14 rounds of MM-UC-MSCs over 12 mo. Her hair began to grow about 3 mo after the first round. The patient was cured, and alopecia universalis did not recur.

MM-UC-MSC transplantation potentially treats patients who suffer from AA and related diseases.

• Alopecia areata
• Umbilical cord-derived mesenchymal stem cells
• Allogenic
• Cell therapy
• Minimal manipulation
• Case report

• colorectal cancer
• mesenchymal stem cell
• oncolytic virus
• p53 mutant tumor
• prodrug activation

• MOST 106-2321-B-039 -003 Minister of Science and Technology

The review summarizes literature data on the role of DNA breaks and DNA repair in the differentiation of pluripotent stem cells (PSC) and connective cell lineages. PSC, including embryonic stem cells (ESC) and induced pluripotent stem cells (iPSC), are rapidly dividing cells with highly active DNA damage response (DDR) mechanisms to ensure the stability and integrity of the DNA. In PSCs, the most common DDR mechanism is error-free homologous recombination (HR) that is primarily active during the S phase of the cell cycle, whereas in quiescent, slow-dividing or non-dividing tissue progenitors and terminally differentiated cells, errorprone non-homologous end joining (NHEJ) mechanism of the double-strand break (DSB) repair is dominating. Thus, it seems that reprogramming and differentiation induce DNA strand breaks in stem cells which itself may trigger the differentiation process. Somatic cell reprogramming to iPSCs is preceded by a transient increase of the DSBs induced presumably by the caspase-dependent DNase or reactive oxygen species. In general, pluripotent stem cells possess stronger DNA repair systems compared to differentiated cells. Nonetheless, during a prolonged cell culture propagation, DNA breaks can accumulate due to the DNA polymerase stalling. Consequently, the DNA damage might trigger the differentiation of stem cells or replicative senescence of somatic cells. The differentiation process per se is often accompanied by a decrease in the DNA repair capacity. Thus, the differentiation might be triggered by DNA breaks, alternatively, the breaks can be a consequence of the decay in the DNA repair capacity of differentiated cells.

• biomaterials
• cell therapy
• hydrogels
• mesenchymal stromal cell
• secretome

• Biocompatible Materials
• Hydrogels
• Mesenchymal Stem Cells

• R01 DE016523 NIDCR NIH HHS

• mesenchymal stem/stromal cells
• single-cell RNA sequencing
• subpopulation
• transcriptomics
• umbilical cord

Human mesenchymal stem cells (hMSCs) promote endogenous tissue regeneration and have become a promising candidate for cell therapy. However, in vitro culture expansion of hMSCs induces a rapid decline of stem cell properties through replicative senescence. Here, we characterize metabolic profiles of hMSCs during expansion. We show that alterations of cellular nicotinamide adenine dinucleotide (NAD + /NADH) redox balance and activity of the Sirtuin (Sirt) family enzymes regulate cellular senescence of hMSCs. Treatment with NAD + precursor nicotinamide increases the intracellular NAD + level and re-balances the NAD + /NADH ratio, with enhanced Sirt-1 activity in hMSCs at high passage, partially restores mitochondrial fitness and rejuvenates senescent hMSCs. By contrast, human fibroblasts exhibit limited senescence as their cellular NAD + /NADH balance is comparatively stable during expansion. These results indicate a potential metabolic and redox connection to replicative senescence in adult stem cells and identify NAD + as a metabolic regulator that distinguishes stem cells from mature cells. This study also suggests potential strategies to maintain cellular homeostasis of hMSCs in clinical applications.

Yuan et al. characterise metabolic profiles of human mesenchymal stem cells (hMSCs) during cell expansion in culture. They find that late passage hMSCs exhibit a NAD + /NADH redox cycle imbalance and that adding NAD + precursor nicotinamide restores mitochondrial fitness and cellular homeostasis in senescent hMSCs indicating a possible route to preserve hMSC homeostasis for therapeutic use.

• Animals
• Bone Marrow Cells/physiology
• Catalase/metabolism
• Cell Culture Techniques
• Cell Proliferation
• Cells, Cultured
• Genomic Instability
• Glutathione/metabolism
• Kidney/cytology
• Kidney/physiology
• Mesenchymal Stem Cell Transplantation
• Mesenchymal Stem Cells/physiology
• Oxidative Stress
• Stem Cells/physiology
• Sus scrofa
• Thiobarbituric Acid Reactive Substances/metabolism

• Cell cycle
• Cell senescence
• DNA damage
• Mesenchymal stem cells

• HNSCC
• MSC
• head and neck cancer
• mesenchymal stem cells
• radiation treatment

Recently, cancer stem cells (CSCs) have been identified as the major cause of both chemotherapy and radiotherapy resistance. Evidence from experimental studies applying both in vitro and in vivo preclinical models suggests that CSCs survive after conventional therapy protocols. Several mechanisms are proposed to be involved in CSC resistance to radiotherapy. Among them, stimulated DNA double-strand break (DSB) repair capacity in association with aldehyde dehydrogenase (ALDH) activity seems to be the most prominent mechanism. However, thus far, the pathway through which ALDH activity stimulates DSB repair is not known. Therefore, in the present study, we investigated the underlying signaling pathway by which ALDH activity stimulates DSB repair and can lead to radioresistance of breast cancer cell lines in vitro. When compared with ALDH-negative cells, ALDH-positive cells presented significantly enhanced cell survival after radiation exposure. This enhanced cell survival was associated with stimulated Nanog, BMI1 and Notch1 protein expression, as well as stimulated Akt activity. By applying overexpression and knockdown approaches, we clearly demonstrated that Nanog expression is associated with enhanced ALDH activity and cellular radioresistance, as well as stimulated DSB repair. Akt and Notch1 targeting abrogated the Nanog-mediated radioresistance and stimulated ALDH activity. Overall, we demonstrate that Nanog signaling induces tumor cell radioresistance and stimulates ALDH activity, most likely through activation of the Notch1 and Akt pathways.

• ALDH1
• Akt
• Nanog
• Notch
• cancer stem cells
• radioresistance

• cell manufacturing
• hydrogels
• mechanotransduction
• mesenchymal stem cells

• R01 DE016523 NIDCR NIH HHS
• R21 AR067469 NIAMS NIH HHS
• S10 OD021601 NIH HHS

• R01 CA174966 NCI NIH HHS
• U.S. Department of Health & Human Services | NIH | National Cancer Institute (NCI)

• ATP-dependent chromatin remodeling
• CSB
• Cockayne syndrome protein
• DNA repair
• PARP1
• base-excision repair (BER)
• chromatin
• oxidative stress
• reactive oxygen species (ROS)
• single-strand DNA repair
• transcription regulation

The stromal vascular fraction (SVF) derived from adipose tissue contains adipose-derived stromal/stem cells (ASC) and can be used for regenerative applications. Thus, a validated protocol for SVF isolation, freezing, and thawing is required to manage product administration. To comply with Good Manufacturing Practice (GMP), fetal bovine serum (FBS), used to expand ASC in vitro, could be replaced by growth factors from platelet concentrates.

Throughout each protocol, GMP-compliant reagents and devices were used. SVF cells were isolated from lipoaspirates by a standardized enzymatic protocol. Cells were cryopreserved in solutions containing different albumin or serum and dimethylsulfoxide (DMSO) concentrations. Before and after cryopreservation, we analyzed: cell viability (by Trypan blue); immunophenotype (by flow cytometry); colony-forming unit-fibroblast (CFU-F) formation; and differentiation potential. ASC, seeded at different densities, were expanded in presence of 10% FBS or 5% supernatant rich in growth factors (SRGF) from platelets. The differentiation potential and cell transformation grade were tested in expanded ASC.

We demonstrated that SVF can be obtained with a consistent yield (about 185 × 10³ cells/ml lipoaspirate) and viability (about 82%). Lipoaspirate manipulation after overnight storage at +4 °C reduced cell viability (−11.6%). The relative abundance of ASC (CD34⁺CD45⁻CD31^–) and endothelial precursors (CD34⁺CD45⁻CD31⁺) in the SVF product was about 59% and 42%, respectively. A period of 2 months cryostorage in autologous serum with added DMSO minimally affected post-thaw SVF cell viability as well as clonogenic and differentiation potentials. Viability was negatively affected when SVF was frozen at a cell concentration below 1.3 × 10⁶ cells/ml. Cell viability was not significantly affected after a freezing period of 1 year.

Independent of seeding density, ASC cultured in 5% SRGF exhibited higher growth rates when compared with 10% FBS. ASC expanded in both media showed unaltered identity (by flow cytometry) and were exempt from genetic lesions. Both 5% SRGF- and 10% FBS-expanded ASC efficiently differentiated to adipocytes, osteocytes, and chondrocytes.

This paper reports a GMP-compliant approach for freezing SVF cells isolated from adipose tissue by a standardized protocol. Moreover, an ASC expansion method in controlled culture conditions and without involvement of animal-derived additives was reported.

• Adipose stem/stromal stem cells
• Adipose tissue
• Advanced therapy medicinal product
• Anchorage independent growth
• CFU-F
• Cell morphology
• Cell viability
• Differentiation potential
• Freezing protocol
• Good manufacturing practice
• Growth rate
• Immunophenotype characterization
• Karyotype
• Stromal vascular fraction