Number Of Senescent Cells Research Articles

Rhoifolin ameliorates osteoarthritis via the Nrf2/NF-κB axis: in vitro and in vivo experiments

Osteoarthritis (OA) is an age-related degenerative disease accompanied by an increasing number of senescent cells and chronic low-grade inflammation. Rhoifolin (ROF) showed considerable inhibition to inflammation, but its role in chondrocyte senescence and OA progress has not been fully characterized. We aimed to evaluate the protective effects of ROF on OA through a series of in vitro and in vivo experiments. The role of ROF in the expression of senescence-associated secretory phenotype (SASP) factors was investigated using RT-qPCR, western blotting, and ELISA. Chondrocyte senescence was assessed by SA-β-gal staining. We applied molecular docking to screen candidate proteins regulated by ROF. Meanwhile, SASP factors and cellular senescence were further assessed after the transfection of Nrf2 siRNA. In the anterior cruciate ligament transection (ACLT) rat model, X-ray, hematoxylin-eosin (HE), and Masson's staining were performed to evaluate the therapeutic effects of ROF on OA. We found that ROF inhibited SASP factors expression and senescence phenotype in IL-1β-treated chondrocytes. Furthermore, ROF suppressed IL-1β-induced activation of the NF-κB pathway cascades. Also, molecular docking and knock-down studies demonstrated that ROF might bind to Nrf2 to suppress the NF-κB pathway. In vivo, ROF ameliorated the OA process in the ACLT rat model. ROF inhibits SASP factors expression and senescence phenotype in chondrocytes and ameliorates the progression of OA via the Nrf2/NF-κB axis, which supports ROF as a potential therapeutic agent for the treatment of OA.

Polymethoxyflavones from Kaempferia parviflora ameliorate skin aging in primary human dermal fibroblasts and ex vivo human skin.

Skin aging is accompanied by an increase in the number of senescent cells, resulting in various pathological outcomes. These include inflammation, impaired barrier function, and susceptibility to skin disorders such as cancer. Kaempferia parviflora (Thai black ginger), a medicinal plant native to Thailand, has been shown to counteract inflammation, cancer, and senescence. This study demonstrates that polymethoxyflavones (5,7-dimethoxyflavone, 5,7,4'-trimethoxyflavone, and 3,5,7,3',4'-pentamethoxyflavone) purified from K. parviflora rhizomes suppressed cellular senescence, reactive oxygen species, and the senescence-associated secretory phenotype in primary human dermal fibroblasts. In addition, they increased tropocollagen synthesis and alleviated free radical-induced cellular and mitochondrial damage. Moreover, the compounds mitigated chronological aging in a human ex vivo skin model by attenuating senescence and restoring expression of essential components of the extracellular matrix, including collagen type I, fibrillin-1, and hyaluronic acid. Finally, we report that polymethoxyflavones enhanced epidermal thickness and epidermal-dermal stability, while blocking age-related inflammation in skin explants. Our findings support the use of polymethoxyflavones from K. parviflora as natural anti-aging agents, highlighting their potential as active ingredients in cosmeceutical and nutraceutical products.

Innovative Treatment for Reducing Senescence and Revitalizing Aging Cells through Gene Silencing

Many studies explore whether harnessing a cellular state known as senescence holds the solution to revitalizing aging tissues. Senescent cells are unique in that they stop cell division but are resistant to apoptosis, a programmed cell death. Senescent cells continue to release chemicals that can trigger inflammation. mTOR inhibitors are currently the only known pharmacological interventions that increase lifespan and revitalize aging tissues in animal models. However, the side effect profiles of mTOR inhibitors are a major cause of concern because mTOR inhibitors inhibit the genes that regulate cell proliferation and immune cell differentiation. To explore a possible solution to the side effects of an mTOR inhibitor, we specifically targeted two mTOR downstream genes, RPS6KB1 (G1 cell cycle inducer) and PPARGC1A (mitochondria energy metabolism regulator) using a small interfering RNA (siRNA) to inhibit gene expression. We hypothesized that inhibiting expression of RPS6KB1 and PPARGC1A would decrease senescence caused by hydrogen peroxide (H2O2). We used human skin cells (Detroit 551) to test our hypothesis and induced senescence by H2O2 treatment. Our result indicates that when both genes were knocked down by siRNA on Detroit 551 cells pretreated with H2O2, the number of senescent cells significantly decreased compared to scrambled siRNA transfected control cells. Our results suggest that inhibiting gene expression of both RPS6KB1 and PPARGC1A may inhibit senescence. Our results may support the development of novel treatments for revitalizing aging tissues and inhibiting inflammation triggered by senescent cells.

Sodium-glucose cotransporter type 2 inhibitors prevent ponatinib-induced endothelial senescence and disfunction: A potential rescue strategy

BackgroundPonatinib (PON), a third-generation tyrosine kinase inhibitor (TKI), has proven cardiovascular toxicity, with no known preventing agents usable to limit such side effect. Sodium-glucose cotransporter type 2 (SGLT2) inhibitors are a new class of glucose-lowering agents, featuring favorable cardiac and vascular effects. AimsWe assessed the effects of the SGLT2 inhibitors empagliflozin (EMPA) and dapagliflozin (DAPA) on human aortic endothelial cells (HAECs) and underlying vasculo-protective mechanisms in an in vitro model of PON-induced endothelial toxicity. Methods and resultsWe exposed HAECs to PON or vehicle (DMSO) in the presence or absence of EMPA (100 and 500 nmol/L) or dapagliflozin (DAPA) for 0–48 h exposure times. Compared with vehicle, incubations of HAECs with PON significantly reduced cell viability (0.56 ± 0.11 vs 0.23 ± 0.05 absorbance units, p < 0.01), increased the number of senescent cells at β-gal-assay (PON 9 ± 4 vs basal DMSO 3 ± 1 β-Gal+ cells/field, p < 0.01), decreased tubulization in Matrigel (PON PON: 6 ± 1 vs basal DMSO 12 ± 1 tubuli number/field, p < 0.05) with a non-statistically significant trend of PON to decrease the number of autophagic cells at immunofluorescence assay and flow cytometry. EMPA reverted the effects of PON on cell viability (E 500 + PON 0.24 ± 0.05 vs PON 0.56 ± 0.11 absorbance units, p < 0.01) and induced autophagy (E 500 7 ± 4.3 vs basal DMSO 2.6 ± 2.3 mean fluorescence vs PON 2.6 ± 2.4 mean fluorescence, p < 0.05). EMPA and DAPA also reversed the effects of PON on cell senescence (E 500 + PON 4 ± 1 and DAPA 100 4 ± 2 vs PON 9 ± 4 β-Gal+ cells/field, p < 0.01) and improved cell tubulization (E 500 + PON 21 ± 3 vs PON 6 ± 1 tubuli number/field, p < 0.05; DAPA 100 + PON 16 ± 2 vs PON 6 ± 1 tubuli number/field, p < 0.05). ConclusionEMPA and DAPA attenuate the vasculo-toxic effect exerted by PON by reverting endothelial cell senescence and dysfunction. These findings support the design of clinical studies exploring the vasculo-protective effects of EMPA or DAPA on PON-induced vascular toxicity.

Abstract 12138: Cell Senescence Associated H3K9me3 Decline Sensitizes Aortic Valve Cell for Osteogenic Response

Introduction: Cell senescence has been identified as an impact etiological factor in cardiovascular diseases, as well as calcific aortic valve disease (CAVD) . In our previous experiment results, We found that a large number of senescent aortic valve interstitial cells (AVIC) accumulated in the calcific aortic valve tissues, especially around the calcification sites. But the role and mechanism of cellular senescence in the development of calcified aortic stenosis have not been clarified. We hypothesized valvular interstitial cell senescence sensitize interstitial cell for osteogenic response through enhancing cell senescence associated epigenetic modification. Methods and Results: Representative immunofluorescence results of the clinical samples show that p16INK4A positive cells in the calcific aortic valve expressed more osteogenic associated factors, such as Runx2 and BMP2. And RAS system hyperactivation seemed to be an impact association with aortic valve calcification. Then we found Angiotensin II (1μM, 72h) chronic irritation induces valve interstitial cell senescence associated secretion phenotype (SASP) and activates osteogenic response through ERK1/2 and NF-κB pathway. ChIP-qPCR results demonstrated H3K9Me3 modifications on the promoters of Runx2, ACTA2(also known as α-SMA), IL-6, IL-8, MCP-1 were significantly decreased in angiotensin II pretreated AVIC. Blockage of angiotensin II and ABT-263 pretreatment inhibited valvular interstitial cell to convert into osteoblast-like phenotype induced by Angiotensin II, as well as attenuating senescence associated secretion phenotype factors yield. Blockage of angiotensin II receptor and p16 INK4A knockdown contributed to recover H3K9me3 modification on the promoters of those gene in angiotensin II pretreated AVIC through attenuating angiotensin II-induced ubiquitination degradation of SUV39H1. Conclusion: p16 INK4A mediated H3K9me3 decline sensitizes aortic valvular interstitial cell for osteogenic response, which impacts on aortic valve homeostasis and aortic valve calcification. Taken together, p16 INK4A positive senescent cells would be the potential target on prevention and treatment of calcific aortic valve disease.

Accumulation of Senescent Neural Cells in Murine Lupus With Depression-Like Behavior.

Neuropsychiatric manifestations targeting the central, peripheral, and autonomic nervous system are common in systemic lupus erythematosus (SLE); collectively, these symptoms are termed neuropsychiatric SLE (NPSLE). Among a wide variety of neuropsychiatric symptoms, depression is observed in about 24-39% of SLE patients. Several cytokines and chemokines have been identified as biomarkers or therapeutic targets of NPSLE; in particular, the levels of type 1 interferons, TNFs, and IL-6 are elevated in SLE patient’s cerebrospinal fluid (CSF), and these factors contribute to the pathology of depression. Here, we show that senescent neural cells accumulate in the hippocampal cornu ammonis 3 (CA3) region in MRL/lpr SLE model mice with depressive behavior. Furthermore, oral administration of fisetin, a senolytic drug, reduced the number of senescent neural cells and reduced depressive behavior in the MRL/lpr mice. In addition, transcription of several senescence and senescence-associated secretory phenotype (SASP) factors in the hippocampal region also decreased after fisetin treatment in the MRL/lpr mice. These results indicate that the accumulation of senescent neural cells in the hippocampus plays a role in NPSLE pathogenesis, and therapies targeting senescent cells may represent a candidate approach to treat NPSLE.

Biomolecular Evaluation of Piceatannol's Effects in Counteracting the Senescence of Mesenchymal Stromal Cells: A New Candidate for Senotherapeutics?

Several investigations on senescence and its causative role in aging have underscored the importance of developing senotherapeutics, a field focused on killing senescent cells and/or preventing their accumulation within tissues. Using polyphenols in counteracting senescence may facilitate the development of senotherapeutics given their presence in the human diet, their confirmed tolerability and absence of severe side effects, and their role in preventing senescence and inducing the death of senescent cells. Against that background, we evaluated the effect of piceatannol, a natural polyphenol, on the senescence of mesenchymal stromal cells (MSCs), which play a key role in the body’s homeostasis. Among our results, piceatannol reduced the number of senescent cells both after genotoxic stress that induced acute senescence and in senescent replicative cultures. Such senotherapeutics activity, moreover, promoted the recovery of cell proliferation and the stemness properties of MSCs. Altogether, our findings demonstrate piceatannol’s effectiveness in counteracting senescence by targeting its associated pathways and detecting and affecting P53-dependent and P53-independent senescence. Our study thus suggests that, given piceatannol’s various mechanisms to accomplish its pleiotropic activities, it may be able to counteract any senescent phenotypes.

Radiation Induced and FACS-Sorted Senescent tdTOMp16+ Cells Upregulate Profibrotic Gene Expression in Mesenchymal Stem Cells (Stromal Cells)

The role of cellular senescence in radiation induced pulmonary fibrosis (RIPF) is unknown. Furthermore, clearance of senescent cells (SCs) could be an effective therapeutic strategy; however, the identification and targeted removal of only senescent cells by senolytic drugs is difficult. We isolated a pure population of radiation induced senescent and non-senescent cells using tdTOMp16+ mice and examined induction of fibrosis in target cells.A stromal cell line was established from explanted tdTOMp16+ mouse bone marrow. The tdTOMp16+ stromal cell line was irradiated to each of several doses by Cesium Gamma Cell dose rate 300 cGy/min including 1.0 Gy, 2.5 Gy, 5 Gy, and 7.5 Gy under growth conditions of either 50% or 100% confluence. The number and percent of senescent cells, by p16 promoter linked to the red fluorochrome biomarker was quantitated. Irradiated red, irradiated non-red, and non-irradiated cells were sorted and cultured as single cells in 96-well plates, then monitored for cell division over four weeks. The expression of senescence markers, p16 and p21, was evaluated by qPCR, and SA-β-gal by fluorogenic substrate. To examine the effect of senescent cells on a target fibrosis-prone C57BL/6 line, sorted irradiated red, irradiated non-red, and non-irradiated cells were placed on the top level of transwells, and after 2 weeks induction of biomarkers of fibrosis including Ctgf, TGF-β, collagen 1a, and collagen 3 was measured in target cells in the lower level by qPCR.After 5 Gy irradiation at 50% cell density, ∼9% of tdTOMp16+ cells became senescent (stably red in color) after 10 days, thus, defining the optimal conditions. Then, in bulk cultures, red cells were sorted by fluorescence automated cell sorter (FACS) to a pure population of red tdTOMp16+ cells. When cultured as single cells, none of 960 irradiated red cells divided after 2 weeks, 137 of these cells remained intact, and assume large and flat morphology. In contrast, 50 of 800 non-red irradiated cells divided, and 104 of 560 non-irradiated control cells divided. There was significant detection of senescent cell markers including: SA-β-gal, p16, and p21 in irradiated sorted red cells compared to irradiated non-red cells (P = 0.0096, P < 0.0001 and P = 0.0001 respectively based on scoring of 1000 cells in triplicate experiments for SA-β-gal and analyses from three independent experiments). Induction of biomarkers of fibrosis Ctgf, TGF-β, collagen 1a, and collagen 3 in C57BL/6 stromal target cells was significant by irradiated red, but not irradiated non-red or non-irradiated cells (P = 0.037, P = 0.14, P = 0.015, and P = 0.0259).The data indicate that sorted irradiated red senescent tdTOMp16 stromal cells can be isolated as a pure non-dividing population and induce fibrotic markers in target cells in non-contact transwell cultures. This unique system can now be used to define the mechanism of senescent cell-induced radiation fibrosis, and the effects of senolytic drugs.

Anti-oncogenic PTEN induces ovarian cancer cell senescence by targeting P21.

Deletion and mutation of phosphatase and tensin homolog deleted on chromosome10 (PTEN) are closely associated with the occurrence of tumors. Tumor suppressor gene PTEN mutation plays an important role in the pathogenesis of ovarian cancer. However, it has been unclear whether it can regulate the senescence of ovarian cancer cells. We speculated that PTEN might inhibit the occurrence and development of ovarian cancer by promoting the expression of P21. We found that the expression of TRIM39 in human ovarian cancer was significantly diminished. In SKOV3 cells treated with naringin, the expression of TRIM39, which binds P21 and inhibits P21 degradation, was significantly elevated. Real‐time polymerase chain reaction (PCR), Western blot, and immunofluorescence were used to detected the expression of PTEN, p21, and TRIM39, β‐galactosidase Staining was used to detect cell senescence, Ki67 staining was used to observe cell proliferation, Trim39 interference or overexpression assay was used to detect its function. We speculated that PTEN might promote SKOV3 cell senescence by increasing TRIM39 expression and decreasing P21 degradation. Furthermore, by interfering with TRIM39 in SKOV3 cells, we found that the expression of P21 was downregulated, and the number of senescent SKOV3 cells decreased. With overexpression of TRIM39 in SKOV3 cells, the expression of P21 was upregulated, and the number of senescent SKOV3 cells increased. When naringin, a PTEN agonist, was added to SKOV3 cells in which TRIM39 protein was interfered with, the expression of P21 was significantly lower than that in the control group, and the number of senescent ovarian cancer cells was significantly diminished. Our results indicated that PTEN maintained the stability of P21 and decreased the degradation of P21 by increasing TRIM39 expression, thus promoting the senescence of SKOV3 cells, and PTEN maintained the stability of p21 and promoted the aging of SKOV3 cells might be a novel therapeutic target for ovarian cancer.

Extracellular matrix and cellular senescence in venous leg ulcers

High prevalence of non-healing chronic wounds contributes to a huge healthcare burden across the world. Early treatment interventions for non-healing wounds are vital. It was previously shown that accumulation of 15% or more of senescent cells in a chronic wound edge is an indicator that the wound is unlikely to heal. However, determining the presence of senescent cells would require invasive procedures such as tissue biopsies to be taken. In this study, we found a strong correlation between decreased collagen area and presence of senescent cells in human chronic wounds i.e. venous leg ulcer (VLU), diabetic foot ulcer (DFU) and pressure ulcer (PRU). We also report that the lowest collagen levels were found in VLU patients less than 60 years of age, with a persistent wound of > 24 months. Elevated levels of senescent cells were also found in VLU of males. Second harmonic imaging of collagen at the edge of chronic wounds with a handheld multiphoton device could be used to predict the number of senescent cells, indicating if the wound is on a healing trajectory or not. Our data support the use of collagen imaging in cutaneous wound assessment for a faster and non-invasive method to predict cellular senescence and determining wound trajectory of healing.

Bradykinin Protects Human Endothelial Progenitor Cells from High-Glucose-Induced Senescence through B2 Receptor-Mediated Activation of the Akt/eNOS Signalling Pathway.

Background Circulating endothelial progenitor cells (EPCs) play important roles in vascular repair. However, the mechanisms of high-glucose- (HG-) induced cord blood EPC senescence and the role of B2 receptor (B2R) remain unknown. Methods Cord blood samples from 26 patients with gestational diabetes mellitus (GDM) and samples from 26 healthy controls were collected. B2R expression on circulating CD34+ cells of cord blood mononuclear cells (CBMCs) was detected using flow cytometry. The plasma concentrations of 8-isoprostaglandin F2α (8-iso-PGF2α) and nitric oxide (NO) were measured. EPCs were treated with HG (40 mM) alone or with bradykinin (BK) (1 nM). The B2R and eNOS small interfering RNAs (siRNAs) and the PI3K antagonist LY294002 were added to block B2R, eNOS, and PI3K separately. To determine the number of senescent cells, senescence-associated β-galactosidase (SA-β-gal) staining was performed. The level of mitochondrial reactive oxygen species (ROS) in EPCs was assessed by Mito-Sox staining. Cell viability was evaluated by Cell Counting Kit-8 (CCK-8) assays. Mitochondrial DNA (mtDNA) copy number and the relative length of telomeres were detected by real time-PCR. The distribution of human telomerase reverse transcriptase (hTERT) in the nucleus, cytosol, and mitochondria of EPCs was detected by immunofluorescence. The expression of B2R, p16, p21, p53, P-Ser473AKT, T-AKT, eNOS, and hTERT was demonstrated by Western blot. Results B2R expression on circulating CD34+ cells of CBMCs was significantly reduced in patients with GDM compared to healthy controls. Furthermore, B2R expression on circulating CD34+ cells of CBMCs was inversely correlated with plasma 8-iso-PGF2α concentrations and positively correlated with plasma NO levels. BK treatment decreased EPC senescence and ROS generation. Furthermore, BK treatment of HG-exposed cells led to elevated P-Ser473AKT and eNOS protein expression compared with HG treatment alone. BK reduced hTERT translocation in HG-induced senescent EPCs. B2R siRNA, eNOS siRNA, and antagonist of the PI3K signalling pathway blocked the protective effects of BK. Conclusion BK, acting through PI3K-AKT-eNOS signalling pathways, reduced hTERT translocation, increased the relative length of telomeres while reducing mtDNA copy number, and finally protected against EPC senescence induced by HG.

Transcription factor EB mediates oxidative stress-induced intervertebral disc degeneration via the NF-κB signaling pathway.

BackgroundIt is well known that the intervertebral disc is aggravated by a significant increase in the number of senescent cells, and oxidative stress (OS) is related to the deterioration of this tissue. Transcription factor EB (TFEB) can protect cells from OS. Accordingly, we investigated whether TFEB can prevent OS in human nucleus pulposus (NP) cells.MethodsFirst, TFEB expression was investigated in human NP tissue samples with different degrees of degeneration. NP cells were treated with different concentrations of hydrogen peroxide (H2O2). The expression of collagen 2, aggrecan, and P65 was detected by quantitative real-time polymerase chain reaction (PCR) and Western blotting. We overexpressed and knocked out the TFEB gene to detect the expression of collagen 2, aggrecan, and P65.ResultsWe found that the expression of TFEB decreased stepwise as the degree of intervertebral disc degeneration (IDD) increased. When the NP cells were treated with H2O2, the expression of TFEB, collagen 2, and aggrecan decreased gradually as H2O2 concentration increased. In addition, the expression of collagen2 and aggrecan increased following TFEB overexpression. However, nuclear factor-kappa B (NF-κB) decreased in NP cells after TFEB overexpression. We also found that the previously low cell viability increased and the high level of apoptosis decreased.ConclusionsThis study suggests that OS is associated with the development of IDD. TFEB mediates OS-induced IDD via the NF-κB signaling pathway. The TFEB gene can potentially be used as a diagnostic biomarker and therapeutic target.

Evaluation of senescent cells in intervertebral discs by lipofuscin staining

Intervertebral disc (IVD) degeneration is considered an important contributor of low back pain, a major age-related disease. Interestingly, an unprecedented high number of senescent cells has been reported in aged and degenerated IVDs, most probably affecting tissue homeostasis. In previous studies classical markers of cellular senescence have been used, such as SA-β-gal staining or p16Ink4a expression. Aim of the presented study was a re-evaluation of the number of senescent IVD cells by using a newly established staining procedure for lipofuscin, based on a Sudan Black-B analogue (GL13), which can be used in fresh, as well as in fixed and embedded tissues. In cultures of senescent rat and human IVD cells both SA-β-gal and GL13 gave similar percentages of senescent cells. Similarly, in fresh tissues from old rats the ratios of senescent cells were high with both detection procedures. Finally, in formalin-fixed and paraffin-embedded tissues from humans, a significant increased number of GL13-positive cells was found in herniated tissues, as compared to apparently normal ones, while similar numbers of p16Ink4a-positive cells were observed. These data confirm the significantly enhanced number of senescent cells in aged and degenerated IVDs, most probably contributing to the degeneration of this tissue.

Tumor Secretome to Adoptive Cellular Immunotherapy: Reduce Me Before I Make You My Partner

Adoptive cellular immunotherapy using chimeric antigen receptor (CAR)-modified T cells and Natural Killer (NK) cells are common immune cell sources administered to treat cancer patients. In detail, whereas CAR-T cells induce outstanding responses in a subset of hematological malignancies, responses are much more deficient in solid tumors. Moreover, NK cells have not shown remarkable results up to date. In general, immune cells present high plasticity to change their activity and phenotype depending on the stimuli they receive from molecules secreted in the tumor microenvironment (TME). Consequently, immune cells will also secrete molecules that will shape the activities of other neighboring immune and tumor cells. Specifically, NK cells can polarize to activities as diverse as angiogenic ones instead of their killer activity. In addition, tumor cell phagocytosis by macrophages, which is required to remove dying tumor cells after the attack of NK cells or CAR-T cells, can be avoided in the TME. In addition, chemotherapy or radiotherapy treatments can induce senescence in tumor cells modifying their secretome to a known as “senescence-associated secretory phenotype” (SASP) that will also impact the immune response. Whereas the SASP initially attracts immune cells to eliminate senescent tumor cells, at high numbers of senescent cells, the SASP becomes detrimental, impacting negatively in the immune response. Last, CAR-T cells are an attractive option to overcome these events. Here, we review how molecules secreted in the TME by either tumor cells or even by immune cells impact the anti-tumor activity of surrounding immune cells.

Extracellular vesicles from mesenchymal stromal cells: Therapeutic perspectives for targeting senescence in osteoarthritis.

Osteoarthritis (OA) is a common age-related disease that correlates with a high number of senescent cells in joint tissues. Senescence has been reported to be one of the main drivers of OA pathogenesis, in particular via the release of senescence-associated secretory phenotype (SASP) factors. SASP factors are secreted as single molecules and/or packaged within extracellular vesicles (EVs), thereby contributing to senescent phenotype dissemination. Targeting senescent cells using senolytics or senomorphics has therefore been tested and improvement of OA-associated features has been reported in murine models. Mesenchymal stromal cells (MSCs) and their derived EVs (MSC-EVs) are promising treatments for OA, exerting pleiotropic functions by producing a variety of factors. However, functions of MSCs and MSC-EVs are affected by aging. In this review, we discuss on the impact of the senescent environment on functions of aged MSC-EVs and on the anti-aging properties of MSC-EVs in the context of OA.

Response to Hypoxia in Cognitive Decline.

Inflammaging, the increase of proinflammatory processes with increasing age, has multiple mechanisms from increasing numbers of senescent cells secreting cytokines to changes in metabolic processes. Alterations of oxygen metabolism with aging, especially decreased levels of O2 with age resulting from endocrine and cardiovascular dysfunction as well as desensitization of cellular response to hypoxia, may exacerbate inflammaging, which in turn creates further oxygen metabolic dysfunction. During aging, decline in levels of erythrocyte 2,3-bisphosphoglycerate (2,3-BPG), BPG mutase, and adenosine A2B receptor, a key adenosine signaling receptor that can augment 2,3-BPG expression, may fail to protect sensitive brain tissue from subtly reduced O2 levels, in turn resulting in increased numbers of activated microglia and secretion of proinflammatory cytokines, ultimately promoting inflammaging and senescence of endothelial cells. Interventions to restore O2 levels directly or via increasing 2,3-BPG may help promote cognitive health in old age, but significant work to quantify the degree of reduced O2 during aging in mammals, and especially humans, needs to be pursued.

Senescent cells exacerbate COVID-19

Coronavirus Cellular senescence is a state elicited in response to stress signals and is associated with a damaging secretory phenotype. The number of senescent cells increases with advanced age and this in turn drives age-related diseases. Camell et al. show that senescent cells have an amplified inflammatory response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) (see the Perspective by Cox and Lord). This response is communicated to nonsenescent cells, suppressing viral defense mechanisms and increasing the expression of viral entry proteins. In old mice infected with a SARS-CoV-2–related virus, treatment with senolytics to reduce the senolytic cell burden reduced mortality and increased antiviral antibodies. Science , abe4832, this issue p. [eabe4832][1]; see also abi4474, p. [281][2] [1]: /lookup/doi/10.1126/science.abe4832 [2]: /lookup/doi/10.1126/science.abi4474

Murine models of Duchenne muscular dystrophy: is there a best model?

Murine models of Duchenne muscular dystrophy: is there a best model?

Cell heterogeneity, rather than the cell storage solution, affects the behavior of mesenchymal stem cells in vitro and in vivo

BackgroundMesenchymal stem cells (MSCs) have to be expanded in vitro to reach a sufficient cell dose for the treatment of various diseases. During the process of expansion, some obstacles remain to be overcome. The purpose of this study was to investigate the effects of storage solutions and heterogeneity on the behavior of MSCs in vitro and in vivo.MethodsUmbilical cord MSCs (UC-MSCs) of similar sizes within normal ranges were suspended in three different storage solutions, phosphate buffer solution, normal saline, and Dulbecco’s modified Eagle medium. Then, the ultrastructure, viability, and safety of these cells were compared. Other two UC-MSC populations of different sizes were categorized based on their mean diameters. The ultrastructure, proliferation, immunosuppression, hepatic differentiation potential, and number of senescent cells were investigated and compared. The survival rates of mice after the infusion of UC-MSCs of different sizes were compared.ResultsFor UC-MSCs suspended in different storage solutions, the cell apoptosis rates, ultrastructure, and survival rates of mice were similar, and no differences were observed. Cells with a diameter of 19.14 ± 4.89 μm were categorized as the larger UC-MSC population, and cells with a diameter of 15.58 ± 3.81 μm were categorized as the smaller population. The mean diameter of the larger UC-MSC population was significantly larger than that of the smaller UC-MSC population (p < 0.01). Smaller UC-MSCs had more powerful proliferation and immunosuppressive potential and a higher nucleus-cytoplasm ratio than those of large UC-MSCs. The number of cells positive for β-galactosidase staining was higher in the larger UC-MSC population than in the smaller UC-MSC population. The survival rates of mice receiving 1 × 106 or 2 × 106 smaller UC-MSCs were 100%, both of which were higher than those of mice receiving the same amounts of larger UC-MSCs (p < 0.01). The cause of mouse death was explored and it was found that some larger UC-MSCs accumulated in the pulmonary capillary in dead mice.ConclusionDifferent storage solutions showed no significant effects on cell behavior, whereas heterogeneity was quite prevalent in MSC populations and might limit cells application. Hence, it is necessary to establish a more precise standardization for culture-expanded MSCs.

N-Acetyl-L-Cysteine Improves Mesenchymal Stem Cell Function in Prolonged Isolated Thrombocytopenia Post-Allotransplant

Background: Prolonged isolated thrombocytopenia (PT), is a serious complication after allogeneic hematopoietic stem cell transplantation (allo-HSCT) and defined as the engraftment of all peripheral blood cell lines other than a platelet (PLT) count ≤20×10E+9/L or dependence on PLT transfusions for more than 60 days after allo-HSCT. Several clinical risk factors have been proposed to be associated with PT after allo-HSCT. However, the underlying mechanisms remain to be elucidated. The processes of megakaryocytopoiesis and thrombocytopoiesis result from the interactions between hematopoietic progenitor cells, cytokines, and bone marrow (BM) stromal cells derived from mesenchymal stem cells (MSCs) or MSCs directly. However, little is known about the quantity and function of BM MSCs and how to improve the impaired BM MSCs in PT patients.AimsTo evaluated the number and function of BM MSCs in patients with PT post-allotransplant. Moreover, to investigate the approach to enhance the number and function of BM MSCs derived from patients with PT and its underlying molecular mechanisms in vitro.MethodsThree cohorts were included: patients with PT (N=83), patients with good graft function (GGF, N=32), defined as persistent successful engraftment after allotransplant, and transplant donors as normal controls (N=15). The number and functions of BM MSCs were evaluated by fibroblasts colony-forming unit (CFU-F) assay, cell proliferation assay and senescence-associated β-galactosidase (SAβ-gal) assay. Reactive oxygen species (ROS) levels and protein expression for p-p38, p38, p-p53, p53 was measured by flow cytometry. The number of megakaryocyte (MK), MK apoptosis, MK ploidy distribution and platelet release values after coculture were assessed by flow cytometry. To further investigate the potential effect for repairing the dysfunctional BM MSCs, N-Acetyl-L-cysteine (NAC,a ROS scavenger), SB203580 (p38 inhibitor) and PFT-a (p53 inhibitor) were administrated to the BM MSCs from PT patients. After 2 days in vitro culture, the percentage of SAβ-positive cells, the intracellular levels of ROS, p-p38, p-p53, the number and the function of MK were evaluated. For evaluation of the efficacy and safety of NAC in PT patients, 20 patients with PT were subsequently enrolled to a single-center pilot study.ResultsHuman BM MSCs were demonstrated as spindle shape and typical immunophenotype of MSCs at day 21 of cultivation among subjects with PT, GGF and normal controls. 7 of the 32 GGF BM and 58 of the 83 PT BM failed to produce any adherent layers within 3 weeks of culture. BM MSCs derived from PT patients expanded more slowly and appeared flattened and larger. Proliferative capacity and CFU-F counts of BM MSCs from PT patients were significantly reduced compared to those of GGF patients and normal controls. Moreover, increased levels of ROS, which was associated with increased number of SAβ-positive cells, were identified in BM MSCs from PT patients. Intracellular p-p38 and p-p53 levels were significantly elevated in PT patients compared to those in GGF patients. Moreover, the percentages of MK were lower in CD34+ cells cocultured with PT-MSCs compared to those observed with GGF-MSCs. CFU-MKs counts were also lower in the PT-MSCs group. After NAC treatment in vitro, the proliferative capacity was increased significantly, whereas the number of senescent cells, the intracellular levels of ROS, p-p38 and p-p53 were reduced markedly in BM MSCs from PT patients. NAC exposure also improved the ability of BM MSCs from PT patients to support megakaryocytopoiesis. The preliminary clinical data of a single-center pilot study showed the complete response, response, and overall response rates at 2 months after NAC treatment (400 mg three times a day) were 10% (2/20), 55% (11/20) and 65% (13/20), respectively, and no apparent drug-associated side effects were observed.Summary/ConclusionIn summary, the current study demonstrated the abnormalities of BM MSCs contribute to the pathogenesis of PT following allo-HSCT. Moreover, BM MSCs derived from PT patients exhibited quantitative and functional improvement following in vitro treatment with NAC. Although these findings require further validation, our in vitro data and preliminary clinical data suggested NAC is an effective and safe treatment option for PT patients after allo-HSCT. DisclosuresNo relevant conflicts of interest to declare.