Exosomal miR-9-5p derived from iPSC-MSCs ameliorates doxorubicin-induced cardiomyopathy by inhibiting cardiomyocyte senescence.

Doxorubicin (DOX) is a chemotherapeutic agent widely used for tumor treatment. Nonetheless its clinical application is heavily limited by its cardiotoxicity. There is accumulated evidence that transplantation of mesenchymal stem cell-derived exosomes (MSC-EXOs) can protect against Dox-induced cardiomyopathy (DIC). This study aimed to examine the cardioprotective effects of EXOs isolated from human induced pluripotent stem cell-derived MSCs (iPSC-MSCs) against DIC and explore the potential mechanisms. EXOs were isolated from the cultural supernatant of human BM-MSCs (BM-MSC-EXOs) and iPSC-MSCs (iPSC-MSC-EXOs) by ultracentrifugation. A mouse model of DIC was induced by intraperitoneal injection of Dox followed by tail vein injection of PBS, BM-MSC-EXOs, or iPSC-MSC-EXOs. Cardiac function, cardiomyocyte senescence and mitochondrial dynamics in each group were assessed. In vitro, neonatal mouse cardiomyocytes (NMCMs) were subjected to Dox and treated with BM-MSC-EXOs or iPSC-MSC-EXOs. The mitochondrial morphology and cellular senescence of NMCMs were examined by Mitotracker staining and senescence-associated-ß-galactosidase assay, respectively. Compared with BM-MSC-EXOs, mice treated with iPSC-MSC-EXOs displayed improved cardiac function and decreased cardiomyocyte mitochondrial fragmentation and senescence. In vitro, iPSC-MSC-EXOs were superior to BM-MSC-EXOs in attenuation of cardiomyocyte mitochondrial fragmentation and senescence caused by DOX. MicroRNA sequencing revealed a higher level of miR-9-5p in iPSC-MSC-EXOs than BM-MSC-EXOs. Mechanistically, iPSC-MSC-EXOs transported miR-9-5p into DOX-treated cardiomyocytes, thereby suppressing cardiomyocyte mitochondrial fragmentation and senescence via regulation of the VPO1/ERK signal pathway. These protective effects and cardioprotection against DIC were largely reversed by knockdown of miR-9-5p in iPSC-MSC-EXOs. Our results showed that miR-9-5p transferred by iPSC-MSC-EXOs protected against DIC by alleviating cardiomyocyte senescence via inhibition of the VPO1/ERK pathway. This study offers new insight into the application of iPSC-MSC-EXOs as a novel therapeutic strategy for DIC treatment.

Downregulation of ALKBH5 rejuvenates aged human mesenchymal stem cells and enhances their therapeutic efficacy in myocardial infarction.

Despite promising results in myocardial infarction (MI), mesenchymal stem cell (MSC)-based therapy is limited by cell senescence. N6-methyladenosine (m6A) messenger RNA methylation has been reported to be closely associated with cell senescence. Nonetheless, its role in the regulation of MSC senescence remains unclear. We examined the role of ALKB homolog 5 (ALKBH5) in regulating MSC senescence and determined whether ALKBH5 downregulation could rejuvenate aged MSCs (AMSCs) to improve their therapeutic efficacy for MI. RNA methylation was determined by m6A dot blotting assay. MSC senescence was evaluated by senescence-associated ß-galactosidase (SA-ß-gal) staining. A mouse model of acute MI was established by ligation of the left anterior decedent coronary artery (LAD). Compared with young MSCs (YMSCs), m6A level was significantly reduced but ALKBH5 was greatly increased in AMSCs. Overexpression of ALKBH5 reduced m6A modification and accelerated YMSC senescence. Conversely, ALKBH5 knockdown increased m6A modifications and alleviated AMSC senescence. Mechanistically, ALKBH5 regulated the m6A modification and stability of CDKN1C mRNA, which further upregulated CDKN1C expression, leading to MSC senescence. CDKN1C overexpression ameliorated the inhibition of cellular senescence of ALKBH5 siRNA-treated AMSCs. More importantly, compared with AMSCs, shALKBH5-AMSCs transplantation provided a superior cardioprotective effect against MI in mice by improving MSC survival and angiogenesis. We determined that ALKBH5 accelerated MSC senescence through m6A modification-dependent stabilization of the CDKN1C transcript, providing a potential target for MSC rejuvenation. ALKBH5 knockdown rejuvenated AMSCs and enhanced cardiac function when transplanted into the mouse heart following infarction.

Prognostic value of APOBEC3A in patients with cervical squamous cell carcinoma in a major urban center in China: a retrospective study.

Background: APOBEC3A (A3A) has been implicated to have vital prognostic value in several common cancers. This study aimed to investigate the prognostic value of A3A expression in cervical squamous cell carcinoma (CESC). Methods: This retrospective study enrolled 59 patients with CESC or cervical squamous intraepithelial neoplasia from January 2014 to January 2017 in Changhai Hospital, Naval Medical University. Then, A3A histoscores (H-scores) using immunohistochemistry (IHC) were analyzed in formalin-fixed paraffin-embedded archival tissue blocks. Moreover, overall survival was analyzed by the Kaplan-Meier method. Results: The H-score of A3A protein expression was relatively higher in CESC than in squamous intraepithelial neoplasia, and the relative expression level of normal cervical tissues was lower than that of cervical squamous intraepithelial neoplasia (P<0.001). Moreover, the H-score of poorly differentiated cases was 6, which was higher than that of moderately differentiated cases (H-score =3), while the H-score of well-differentiated cases was 2, which was lower than that of moderately differentiated cases. Moreover, patients in the A3A low expression group had higher overall survival rates by prognostic analysis (P=0.027). Conclusions: A3A protein expression was increased during CESC progression. Moreover, A3A expression was tightly related to poor prognosis in CESC. Thus, these results showed that A3A overexpression may provide a marker for poor prognosis in CESC.

APOBEC3A suppresses cervical cancer via apoptosis.

Background: Family members of Apolipoprotein B mRNA-editing enzyme catalytic 3 (APOBEC3) play critical roles in cancer evolution and development. However, the role of APOBEC3A in cervical cancer remains to be clarified. Methods: We used bioinformatics to investigate APOBEC3A expression and outcomes using The Cancer Genome Atlas (TCGA)-cervical squamous cell carcinoma and endocervical adenocarcinoma (CESC) dataset, GTEx, and GSE7803. Immunohistochemistry was then used to identify APOBEC3A's expression pattern. We performed Cell Counting Kit-8, wound-healing, Transwell, and flow cytometry assays to measure proliferation, migration, invasion, and apoptosis, respectively, using the SiHa and HeLa cell lines transfected with APOBEC3A. BALB/c nude mice were used to investigate the effects of APOBEC3A in vivo. The phosphorylated gamma-H2AX staining assay was applied to measure DNA damage. RNA sequencing (RNA-Seq) was applied to explore APOBEC3A-related signaling pathways. Results: APOBEC3A was more significantly expressed in cancer tissues than in adjacent normal tissues. Higher expression of APOBEC3A was associated with better outcomes in TCGA-CESC and GTEx. Immunohistochemistry showed that the expression of APOBEC3A was significantly higher in cancer tissues than in normal tissues. Transfection experiments showed that APOBEC3A inhibited proliferation, upregulated S-phase cells, inhibited migration and invasion, induced DNA damage, and promoted apoptosis. Overexpression of APOBEC3A inhibited tumor formation in the mouse model. RNA-seq analysis showed that ectopic expression of APOBEC3A inhibited several cancer-associated signaling pathways. Conclusions: APOBEC3A is significantly upregulated in cervical cancer, and higher expression of APOBEC3A is associated with better outcomes. APOBEC3A is a tumor suppressor whose overexpression induces apoptosis in cervical cancer.

Xiaotan Sanjie recipe, a compound Chinese herbal medicine, inhibits gastric cancer metastasis by regulating GnT-V-mediated E-cadherin glycosylation.

OBJECTIVE: Xiaotan Sanjie recipe (XTSJ), a Chinese herbal compound medicine, exerts a significant inhibitory effect on gastric cancer (GC) metastasis. This work investigated the mechanism underlying the XTSJ-mediated inhibition of GC metastasis. METHODS: The effect of XTSJ on GC metastasis and the associated mechanism were investigated in vitro, using GC cell lines, and in vivo, using a GC mouse model, by focusing on the expression of Glc-N-Ac-transferase V (GnT-V; encoded by MGAT5). RESULTS: The migration and invasion ability of GC cells decreased significantly after XTSJ administration, which confirmed the efficacy of XTSJ in treating GC in vitro. XTSJ increased the accumulation of E-cadherin at junctions between GC cells, which was reversed by MGAT5 overexpression. XTSJ administration and MGAT5 knockdown alleviated the structural abnormality of the cell-cell junctions, while MGAT5 overexpression had the opposite effect. MGAT5 knockdown and XTSJ treatment also significantly increased the accumulation of proteins associated with the E-cadherin-mediated adherens junction complex. Furthermore, the expression of MGAT5 was significantly lower in the lungs of BGC-823-MGAT5 + XTSJ mice than in those of BGC-823-MGAT5 + solvent mice, indicating that the ability of gastric tumors to metastasize to the lung was decreased in vivo following XTSJ treatment. CONCLUSION: XTSJ prevented GC metastasis by inhibiting the GnT-V-mediated E-cadherin glycosylation and promoting the E-cadherin accumulation at cell-cell junctions. Please cite this article as: Huang N, He HW, He YY, Gu W, Xu MJ, Liu L. Xiaotan Sanjie recipe, a compound Chinese herbal medicine, inhibits gastric cancer metastasis by regulating GnT-V-mediated E-cadherin glycosylation. J Integr Med. 2023; 21(6): 561-574.

Moderate altitude exposure impacts host fasting blood glucose and serum metabolome by regulation of the intestinal flora.

Moderate altitude exposure has shown beneficial effects on diabetes incidence but the underlying mechanisms are not understood. Our study aimed to investigate how the human gut microbiome impacted the serum metabolome and associated with glucose homeostasis in healthy Chinese individuals upon moderate-altitude exposure. Faecal microbiome composition was assessed using shotgun metagenomic sequencing. Serum metabolome was acquired by untargeted metabolomics technology, and amino acids (AAs) and propionic acid in serum were quantified by targeted metabolomics technology. The results indicated that the moderate-altitude exposed individuals presented lowered fasting blood glucose (FBG) and propionic acid, increased circulating L-Glutamine but decreased L-Glutamate and L-Valine, which correlated with enriched Bacteroidetes and decreased Proteobacteria. Additionally, the silico causality associations among gut microbiota, serum metabolome and host FBG were analyzed by mediation analysis. It showed that increased Bacteroides ovatus (B. ovatus) and decreased Escherichia coli (E. coli) were identified as the main antagonistic species driving the association between L-Glutamate and FBG in silico causality. Furthermore, the high-fat diet (HFD) fed mice subjected to faecal microbiota transplantation (FMT) were applied to validate the cause-in-fact effects of gut microbiota on the beneficial glucose response. We found that microbiome in the moderate-altitude exposed donor could predict the extent of the FBG response in recipient mice, which showed lowered FBG, L-Glutamate and Firmicutes/Bacteroidetes ratio. Our findings suggest that moderate-altitude exposure targeting gut microbiota and circulating metabolome, may pave novel avenues to counter dysglycemia.

Effects of polymer terminal group inside micelle core on paclitaxel loading promoting and burst release suppressing.

Background: Paclitaxel (PTX) is widely used in the treatment of advanced esophageal and gastric cancer. Polymeric micelles can improve the drug-loading efficiency of PTX. However, the end groups on the amphiphilic blocks affect the drug-loading efficiency and the release kinetics of polymeric micelles. Therefore, there is an urgent need to disclose the tailoring of the core-/shell-forming terminal groups. Methods: Different from the conventional block copolymer synthesis in the reversible addition-fragmentation chain-transfer polymerization, which has a hydrophilic end group on the core-forming blocks, an alternative monomer addition method was applied to tune and obtain two block copolymers with symmetrical and similar block length PBMAn-b-PNAMm [PNAM, poly(N-acryloylmorpholine); PBMA, poly(n-butyl methacrylate)] but distinct end groups on the hydrophobic core-forming blocks, that is, HOOC-PBMA-PNAM-Phen and HOOC-PNAM-PBMA-Phen. The chemical structure of the resulting copolymers was elucidated by proton nuclear magnetic resonance spectroscopy and differential scanning calorimetry. The spherical morphology revealed by transmission electron microscopy and the uniform particle size revealed by dynamic light scattering analysis clearly confirmed the successful preparation of a PTX-polymeric micelle complex. Results: The particle sizes of HOOC-PBMA-PNAM-Phen and HOOC-PNAM-PBMA-Phen were about 40 and 235 nm respectively. The PTX loading efficiency of HOOC-PBMA-PNAM-Phen was much lower than that of HOOC-PNAM-PBMA-Phen. The PTX release from HOOC-PBMA-PNAM-Phen was much slower than that of HOOC-PNAM-PBMA-Phen. The polymers had glass transition temperature (Tg) values of 70.24 and 74.22 ℃, which was from the HOOC-PBMA-PNAM-Phen and HOOC-PNAM-PBMA-Phen micelles, respectively. The systematic study on the PTX loading and releasing profile disclosed that, compared with the HOOC-PBMA-PNAM-Phen, the micelles with Phen group on the hydrophobic block (HOOC-PNAM-PBMA-Phen) enhanced drug loading and prolonged drug release but with a larger particle size. Conclusions: The results indicated that the hydrophobic end group Phen on the core-forming blocks can promote hydrophobic drug loading and suppress burst release.

[Mechanism of Notch3 signaling pathway regulating the differentiation of aortic dissection vascular stem cells into smooth muscle cells].

OBJECTIVE: To explore whether the differentiation of vascular stem cells (VSC) into smooth muscle cells (SMC) in aortic dissection (AD) is dysregulated, and to verify the role of Notch3 pathway in this process. METHODS: Aortic tissues were obtained from AD patients undergoing aortic vascular replacement and heart transplant donors at Department of Cardiovascular Surgery, Guangdong Provincial People's Hospital Affiliated to Southern Medical University. VSC were isolated by enzymatic digestion and c-kit immunomagnetic beads. The cells were divided into normal donor-derived VSC group (Ctrl-VSC group) and AD-derived VSC group (AD-VSC group). The presence of VSC in the aortic adventitia was detected by immunohistochemical staining, and VSC was identified by stem cell function identification kit. The differentiation model of VSC into SMC established in vitro was induced by transforming growth factor-ß1 (10 µg/L) for 7 days. They were divided into normal donor VSC-SMC group (Ctrl-VSC-SMC group), AD VSC-SMC group (AD-VSC-SMC group) and AD VSC-SMC+Notch3 inhibitor DAPT group (AD-VSC-SMC+DAPT group,DAPT 20 µmol/L was added during differentiation induction). The expression of contractile marker Calponin 1 (CNN1) in SMC derived from aortic media and VSC were detected by immunofluorescence staining. The protein expressions of contractile markers α-smooth muscle actin (α-SMA), CNN1 as well as Notch3 intracellular domain (NICD3) in SMC derived from aortic media and VSC were detected by Western blotting. RESULTS: Immunohistochemical staining showed there was a population of c-kit-positive VSC in the adventitia of aortic vessels, and VSC from both normal donors and AD patients had the ability to differentiate into adipocytes and chondrocytes. Compared with normal donor vascular tissue, the expressions of SMC markers α-SMA and CNN1 of tunica media contraction in AD were down-regulated (α-SMA/ß-actin: 0.40±0.12 vs. 1.00±0.11, CNN1/ß-actin: 0.78±0.07 vs. 1.00±0.14, both P < 0.05), while the protein expression of NICD3 was up-regulated (NICD3/GAPDH: 2.22±0.57 vs. 1.00±0.15, P < 0.05). Compared with Ctrl-VSC-SMC group, the expressions of contractile SMC markers α-SMA and CNN1 were down-regulated in AD-VSC-SMC group (α-SMA/ß-actin: 0.35±0.13 vs. 1.00±0.20, CNN1/ß-actin: 0.78±0.06 vs. 1.00±0.07, both P < 0.05), the protein expression of NICD3 was up-regulated (NICD3/GAPDH: 22.32±1.22 vs. 1.00±0.06, P < 0.01). Compared with AD-VSC-SMC group, the expressions of contractile SMC markers α-SMA, CNN1 were up-regulated in AD-VSC-SMC+DAPT group (α-SMA/ß-actin: 1.70±0.07 vs. 1.00±0.15, CNN1/ß-actin: 1.62±0.03 vs. 1.00±0.02, both P < 0.05). CONCLUSIONS: Dysregulation of VSC differentiation into SMC occurs in AD, while inhibition of Notch3 pathway activation can restore the expression of contractile proteins in VSC-derived SMC in AD.

The mortalities of female-specific cancers in China and other countries with distinct socioeconomic statuses: A longitudinal study.

INTRODUCTION: Female-specific cancers seriously affect physical and psychological health of women worldwide. OBJECTIVES: We aimed to elucidate trends in the age-standardized mortality rates (ASMRs) of breast cancer, cervical cancer, uterine cancer, and ovarian cancer in female populations with different socioeconomic statuses in China and in countries with different Human Development Index (HDI). METHODS: A longitudinal study was performed using the data of cancer death in China and other 39 countries. The mortality rates were standardized with the Segi's world population. Trends in the mortalities were exhibited by estimated annual percentage change (EAPC). Pearson correlation was used to assess the association between EAPC and HDI. RESULTS: In mainland China, female breast cancer, cervical cancer, uterine cancer, and ovarian cancer accounted for 6.60 %, 4.21 %, 2.50 %, and 2.02 % of cancer death (n = 1,314,040) in women with 1,220,251,032 person-years, respectively. The ASMRs of cervical cancer (EAPC = 3.87 %, P < 0.001) and ovarian cancer (EAPC = 1.81 %, P < 0.001) increased, that of female breast cancer unchanged, whereas that of uterine cancer was extremely higher and rapidly decreased (EAPC =  - 7.65 %, P < 0.001), during 2004-2019. The ASMRs of female breast and ovarian cancers were higher in urban and developed regions than in rural and undeveloped regions, in contrast to cervical and uterine cancers. The ASMRs of female breast and ovarian cancers were lower in China than in other countries, in contrast to uterine cancer. The ASMR of cervical cancer decreased, that of uterine cancer increased, in other countries during 2004-2017. EAPCs for the ASMRs of breast and ovarian cancers were inversely correlated to HDI. CONCLUSION: The ASMRs of cervical and ovarian cancers increased, in contrast to uterine cancer, in China during socioeconomic transition. Trends in the ASMRs of breast and ovarian cancers were inversely associated with HDI. These data help control female-specific cancers.

Reversibly Migratable Fluorescent Probe for Precise and Dynamic Evaluation of Cell Mitochondrial Membrane Potentials.

The mitochondrial membrane potential (MMP, ΔΨmito) provides the charge gradient required for mitochondrial functions and is a key indicator of cellular health. The changes in MMP are closely related to diseases and the monitoring of MMP is thus vital for pathological study and drug development. However, most of the current fluorescent probes for MMP rely solely on the cell fluorescence intensity and are thus restricted by poor photostability, rendering them not suitable for long-term dynamic monitoring of MMP. Herein, an MMP-responsive fluorescent probe pyrrolyl quinolinium (PQ) which is capable of reversible migration between mitochondria and nucleolus is developed and demonstrated for dynamic evaluation of MMP. The fluorescence of PQ translocates from mitochondria to nucleoli when MMP decreases due to the intrinsic RNA-specificity and more importantly, the translocation is reversible. The cytoplasm to nucleolus fluorescence intensity ratio is positively correlated with MMP so that this method avoids the negative influence of photostability and imaging parameters. Various situations of MMP can be monitored in real time even without controls. Additionally, long-term dynamic evaluation of MMP is demonstrated for HeLa cells using PQ in oxidative environment. This study is expected to give impetus to the development of mitochondria-related disease diagnosis and drug screening.

Tri-functional SERS nanoplatform with tunable plasmonic property for synergistic antibacterial activity and antibacterial process monitoring.

Strategies integrating synergistic high-efficiency bacterial killing and antibacterial process monitoring capability are desirable. Herein, a tri-functional surface-enhanced Raman spectroscopy (SERS) nanoplatform, namely 4-mercaptobenzoic acid-encoded gold nanorods@silver coated with a layer of bovine serum albumin (AuNRs@Ag@4-MBA@BSA), with excellent biocompatibility, stability, tunable plasmonic property and activatable photothermal effect is introduced for Ag+/photothermal therapy (PTT) synergistic antibacterial activity and antibacterial process monitoring. An exogenous etchant is used to controllably model the physiological process of metallic silver biodegradation. Ag shell etching causes the surface plasmon resonance band of SERS nanotags to red-shift to near-infrared region, activates the photothermal conversion capability, and triggers PTT, which in turn accelerates Ag shell etching. The antibacterial rates for Staphylococcus aureus and Escherichia coli after 10 min treatment can achieve 99.5% and 99.9%, respectively. Furthermore, the near-field effect and ultrasensitive property render the SERS intensity decrease ratio is dependent on Ag shell etching as well as temperature rising and thus relevant to antibacterial activity. We have demonstrated a strong correlation between SERS signal and antibacterial effect, and have verified the possibility of antibacterial process monitoring in vitro using SERS-based methodology. We envision that our integrated strategy being used for in vivo high-efficiency bacterial killing and antibacterial process monitoring.

MicroRNA-199a-5p aggravates angiotensin II-induced vascular smooth muscle cell senescence by targeting Sirtuin-1 in abdominal aortic aneurysm.

Vascular smooth muscle cells (VSMCs) senescence contributes to abdominal aortic aneurysm (AAA) formation although the underlying mechanisms remain unclear. This study aimed to investigate the role of miR-199a-5p in regulating VSMC senescence in AAA. VSMC senescence was determined by a senescence-associated ß-galactosidase (SA-ß-gal) assay. RT-PCR and Western blotting were performed to measure miRNA and protein level, respectively. The generation of reactive oxygen species (ROS) was evaluated by H2DCFDA staining. Dual-luciferase reporter assay was used to validate the target gene of miR-199a-5p. VSMCs exhibited increased senescence in AAA tissue relative to healthy aortic tissue from control donors. Compared with VSMCs isolated from control donors (control-VSMCs), those derived from patients with AAA (AAA-VSMCs) exhibited increased cellular senescence and ROS production. Angiotensin II (Ang II) induced VSMC senescence by promoting ROS generation. The level of miR-199a-5p expression was upregulated in the plasma from AAA patients and Ang II-treated VSMCs. Mechanistically, Ang II treatment significantly elevated miR-199a-5p level, thereby stimulating ROS generation by repressing Sirt1 and consequent VSMC senescence. Nevertheless, Ang II-induced VSMC senescence was partially attenuated by a miR-199a-5p inhibitor or Sirt1 activator. Our study revealed that miR-199a-5p aggravates Ang II-induced VSMC senescence by targeting Sirt1 and that miR-199a-5p is a potential therapeutic target for AAA.

miR-155-5p inhibition rejuvenates aged mesenchymal stem cells and enhances cardioprotection following infarction.

Aging impairs the functions of human mesenchymal stem cells (MSCs), thereby severely reducing their beneficial effects on myocardial infarction (MI). MicroRNAs (miRNAs) play crucial roles in regulating the senescence of MSCs; however, the underlying mechanisms remain unclear. Here, we investigated the significance of miR-155-5p in regulating MSC senescence and whether inhibition of miR-155-5p could rejuvenate aged MSCs (AMSCs) to enhance their therapeutic efficacy for MI. Young MSCs (YMSCs) and AMSCs were isolated from young and aged donors, respectively. The cellular senescence of MSCs was evaluated by senescence-associated ß-galactosidase (SA-ß-gal) staining. Compared with YMSCs, AMSCs exhibited increased cellular senescence as evidenced by increased SA-ß-gal activity and decreased proliferative capacity and paracrine effects. The expression of miR-155-5p was much higher in both serum and MSCs from aged donors than young donors. Upregulation of miR-155-5p in YMSCs led to increased cellular senescence, whereas downregulation of miR-155-5p decreased AMSC senescence. Mechanistically, miR-155-5p inhibited mitochondrial fission and increased mitochondrial fusion in MSCs via the AMPK signaling pathway, thereby resulting in cellular senescence by repressing the expression of Cab39. These effects were partially reversed by treatment with AMPK activator or mitofusin2-specific siRNA (Mfn2-siRNA). By enhancing angiogenesis and promoting cell survival, transplantation of anti-miR-155-5p-AMSCs led to improved cardiac function in an aged mouse model of MI compared with transplantation of AMSCs. In summary, our study shows that miR-155-5p mediates MSC senescence by regulating the Cab39/AMPK signaling pathway and miR-155-5p is a novel target to rejuvenate AMSCs and enhance their cardioprotective effects.

Exosomes from mesenchymal stem cells overexpressing MIF enhance myocardial repair.

Accumulating evidence has shown that mesenchymal stem cell (MSC)-derived exosomes (exo) mediate cardiac repair following myocardial infarction (MI). Macrophage migration inhibitory factor (MIF), a proinflammatory cytokine, plays a critical role in regulating cell homeostasis. This study aimed to investigate the cardioprotective effects of exo secreted from bone marrow-MSCs (BM-MSCs) overexpressing MIF in a rat model of MI. MIF plasmid was transducted in BM-MSCs. Exo were isolated from the supernatants of BM-MSCs and MIF-BM-MSCs, respectively. The morphology of mitochondria in neonatal mice cardiomyocytes (NRCMs) was determined by MitoTracker staining. The apoptosis of NRCMs was examined by deoxynucleotidyl transferase-mediated dUTP nick end-labeling. BM-MSC-exo and MIF-BM-MSC-exo were intramuscularly injected into the peri-infarct region in a rat model of MI. The heart function of rats was assessed by echocardiography. The expression of MIF was greatly enhanced in MIF-BM-MSCs compared with BM-MSCs. Both BM-MSC-exo and MIF-BM-MSC-exo expressed CD63 and CD81. NRCMs treated with MIF-BM-MSC-exo exhibited less mitochondrial fragmentation and cell apoptosis under hypoxia/serum deprivation (H/SD) challenge than those treated with BM-MSC-exo via activating adenosine 5'-monophosphate-activated protein kinase signaling. Moreover, these effects were partially abrogated by Compound C. Injection of BM-MSC-exo or MIF-BM-MSC-exo greatly restored heart function in a rat model of MI. Compared with BM-MSC-exo, injection of MIF-BM-MSC-exo was associated with enhanced heart function, reduced heart remodeling, less cardiomyocyte mitochondrial fragmentation, reactive oxygen species generation, and apoptosis. Our study reveals a new mechanism of MIF-BM-MSC-exo-based therapy for MI and provides a novel strategy for cardiovascular disease treatment.

Haemin pre-treatment augments the cardiac protection of mesenchymal stem cells by inhibiting mitochondrial fission and improving survival.

The cardiac protection of mesenchymal stem cell (MSC) transplantation for myocardial infarction (MI) is largely hampered by low cell survival. Haem oxygenase 1 (HO-1) plays a critical role in regulation of cell survival under many stress conditions. This study aimed to investigate whether pre-treatment with haemin, a potent HO-1 inducer, would promote the survival of MSCs under serum deprivation and hypoxia (SD/H) and enhance the cardioprotective effects of MSCs in MI. Bone marrow (BM)-MSCs were pretreated with or without haemin and then exposed to SD/H. The mitochondrial morphology of MSCs was determined by MitoTracker staining. BM-MSCs and haemin-pretreated BM-MSCs were transplanted into the peri-infarct region in MI mice. SD/H induced mitochondrial fragmentation, as shown by increased mitochondrial fission and apoptosis of BM-MSCs. Pre-treatment with haemin greatly inhibited SD/H-induced mitochondrial fragmentation and apoptosis of BM-MSCs. These effects were partially abrogated by knocking down HO-1. At 4 weeks after transplantation, compared with BM-MSCs, haemin-pretreated BM-MSCs had greatly improved the heart function of mice with MI. These cardioprotective effects were associated with increased cell survival, decreased cardiomyocytes apoptosis and enhanced angiogenesis. Collectively, our study identifies haemin as a regulator of MSC survival and suggests a novel strategy for improving MSC-based therapy for MI.

Transfer of mitochondria from mesenchymal stem cells derived from induced pluripotent stem cells attenuates hypoxia-ischemia-induced mitochondrial dysfunction in PC12 cells.

Mitochondrial dysfunction in neurons has been implicated in hypoxia-ischemia-induced brain injury. Although mesenchymal stem cell therapy has emerged as a novel treatment for this pathology, the mechanisms are not fully understood. To address this issue, we first co-cultured 1.5 × 105 PC12 cells with mesenchymal stem cells that were derived from induced pluripotent stem cells at a ratio of 1:1, and then intervened with cobalt chloride (CoCl2) for 24 hours. Reactive oxygen species in PC12 cells was measured by Mito-sox. Mitochondrial membrane potential (?Ψm) in PC12 cells was determined by JC-1 staining. Apoptosis of PC12 cells was detected by terminal deoxynucleotidal transferase-mediated dUTP nick end-labeling staining. Mitochondrial morphology in PC12 cells was examined by transmission electron microscopy. Transfer of mitochondria from the mesenchymal stem cells derived from induced pluripotent stem cells to damaged PC12 cells was measured by flow cytometry. Mesenchymal stem cells were induced from pluripotent stem cells by lentivirus infection containing green fluorescent protein in mitochondria. Then they were co-cultured with PC12 cells in Transwell chambers and treated with CoCl2 for 24 hours to detect adenosine triphosphate level in PC12 cells. CoCl2-induced PC12 cell damage was dose-dependent. Co-culture with mesenchymal stem cells significantly reduced apoptosis and restored ?Ψm in the injured PC12 cells under CoCl2 challenge. Co-culture with mesenchymal stem cells ameliorated mitochondrial swelling, the disappearance of cristae, and chromatin margination in the injured PC12 cells. After direct co-culture, mitochondrial transfer from the mesenchymal stem cells stem cells to PC12 cells was detected via formed tunneling nanotubes between these two types of cells. The transfer efficiency was greatly enhanced in the presence of CoCl2. More importantly, inhibition of tunneling nanotubes partially abrogated the beneficial effects of mesenchymal stem cells on CoCl2-induced PC12 cell injury. Mesenchymal stem cells reduced CoCl2-induced PC12 cell injury and these effects were in part due to efficacious mitochondrial transfer.

Adipose-Derived Mesenchymal Stem Cells Isolated from Patients with Abdominal Aortic Aneurysm Exhibit Senescence Phenomena.

Mesenchymal stem cells (MSCs) have shown beneficial effects in the treatment of abdominal aortic aneurysm (AAA). Nonetheless, the biological properties of adipose-derived MSCs (ASCs) from patients with AAA (AAA-ASCs) remain unclear. This study is aimed at investigating the properties of cell phenotype and function of AAA-ASCs compared with ASCs from age-matched healthy donors (H-ASCs). H-ASCs and AAA-ASCs were studied for cell phenotype, differentiation capacity, senescence, and mitochondrial and autophagic functions. Cellular senescence was examined by senescence-associated ß-galactosidase (SA-ß-gal) staining. Mitochondrial morphology was determined by MitoTracker staining. Despite the similar surface markers of AAA-ASCs and H-ASCs, AAA-ASCs exhibited altered multidifferentiation potential. Compared with H-ASCs, AAA-ASCs displayed enhanced senescence manifested by increased SA-ß-gal activity and decreased proliferation and migration ability. Furthermore, AAA-ASCs showed increased mitochondrial fusion, reactive oxygen species (ROS) production, and decreased mitochondrial membrane potential. In addition, AAA-ASCs exhibited decreased autophagy level, upregulation of IL-6 and TNF-α secretion, and downregulation of IL-10 secretion compared with H-ASCs. Nonetheless, treatment of AAA-ASCs with rapamycin (an autophagy activator) dramatically reduced secretion of IL-6 and TNF-α and enhanced secretion of IL-10. In conclusion, our study showed that AAA-ASCs exhibit senescence phenomena and decreased cell function. Understanding the specific alterations in AAA-ASCs will help explore novel strategies to restore cell function for AAA treatment.

Macrophage migration inhibitory factor rejuvenates aged human mesenchymal stem cells and improves myocardial repair.

The beneficial functions of mesenchymal stem cells (MSCs) decline with age, limiting their therapeutic efficacy for myocardial infarction (MI). Macrophage migration inhibitory factor (MIF) promotes cell proliferation and survival. We investigated whether MIF overexpression could rejuvenate aged MSCs and increase their therapeutic efficacy in MI. Young and aged MSCs were isolated from the bone marrow of young and aged donors. Young MSCs, aged MSCs, and MIF-overexpressing aged MSCs were transplanted into the peri-infarct region in a rat MI model. Aged MSCs exhibited a lower proliferative capacity, lower MIF level, greater cell size, greater senescence-associated-ß-galactosidase activity, and weaker paracrine effects than young MSCs. Knocking down MIF in young MSCs induced cellular senescence, whereas overexpressing MIF in aged MSCs reduced cellular senescence. MIF rejuvenated aged MSCs by activating autophagy, an effect largely reversed by the autophagy inhibitor 3-methyladenine. MIF-overexpressing aged MSCs induced angiogenesis and prevented cardiomyocyte apoptosis to a greater extent than aged MSCs, and had improved heart function and cell survival more effectively than aged MSCs four weeks after MI. Thus, MIF rejuvenated aged MSCs by activating autophagy and enhanced their therapeutic efficacy in MI, suggesting a novel MSC-based therapeutic strategy for cardiovascular diseases in the aged population.

Soluble Nogo receptor 1 fusion protein protects neural progenitor cells in rats with ischemic stroke.

Soluble Nogo66 receptor-Fc protein (sNgR-Fc) enhances axonal regeneration following central nervous system injury. However, the underlying mechanisms remain unclear. In this study, we investigated the effects of sNgR-Fc on the proliferation and differentiation of neural progenitor cells. The photothrombotic cortical injury model of ischemic stroke was produced in the parietal cortex of Sprague-Dawley rats. The rats with photothrombotic cortical injury were randomized to receive infusion of 400 µg/kg sNgR-Fc (sNgR-Fc group) or an equal volume of phosphate-buffered saline (photothrombotic cortical injury group) into the lateral ventricle for 3 days. The effects of sNgR-Fc on the proliferation and differentiation of endogenous neural progenitor cells were examined using BrdU staining. Neurological function was evaluated with the Morris water maze test. To further examine the effects of sNgR-Fc treatment on neural progenitor cells, photothrombotic cortical injury was produced in another group of rats that received transplantation of neural progenitor cells from the hippocampus of embryonic Sprague-Dawley rats. The animals were then given an infusion of phosphate-buffered saline (neural progenitor cells group) or sNgR-Fc (sNgR-Fc + neural progenitor cells group) into the lateral ventricle for 3 days. sNgR-Fc enhanced the proliferation of cultured neural progenitor cells in vitro as well as that of endogenous neural progenitor cells in vivo, compared with phosphate-buffered saline, and it also induced the differentiation of neural progenitor cells into neurons. Compared with the photothrombotic cortical injury group, escape latency in the Morris water maze and neurological severity score were greatly reduced, and distance traveled in the target quadrant was considerably increased in the sNgR-Fc group, indicating a substantial improvement in neurological function. Furthermore, compared with phosphate-buffered saline infusion, sNgR-Fc infusion strikingly improved the survival and differentiation of grafted neural progenitor cells. Our findings show that sNgR-Fc regulates neural progenitor cell proliferation, migration and differentiation. Therefore, sNgR-Fc is a potential novel therapy for stroke and neurodegenerative diseases, The protocols were approved by the Committee on the Use of Live Animals in Teaching and Research of the University of Hong Kong (approval No. 4560-17) in November, 2015.

TGF-ß mediates aortic smooth muscle cell senescence in Marfan syndrome.

Formation of aortic aneurysms as a consequence of augmented transforming growth factor ß (TGF-ß) signaling and vascular smooth muscle cell (VSMC) dysfunction is a potentially lethal complication of Marfan syndrome (MFS). Here, we examined VSMC senescence in patients with MFS and explored the potential mechanisms that link VSMC senescence and TGF-ß. Tissue was harvested from the ascending aorta of control donors and MFS patients, and VSMCs were isolated. Senescence-associated ß-galactosidase (SA-ß-gal) activity and expression of senescence-related proteins (p53, p21) were significantly higher in aneurysmal tissue from MFS patients than in healthy aortic tissue from control donors. Compared to control-VSMCs, MFS-VSMCs were larger with higher levels of both SA-ß-gal activity and mitochondrial reactive oxygen species (ROS). In addition, TGF-ß1 levels were much higher in MFS- than control-VSMCs. TGF-ß1 induced VSMC senescence through excessive ROS generation. This effect was suppressed by Mito-tempo, a mitochondria-targeted antioxidant, or SC-514, a NF-κB inhibitor. This suggests TGF-ß1 induces VSMC senescence through ROS-mediated activation of NF-κB signaling. It thus appears that a TGF-ß1/ROS/NF-κB axis may mediate VSMC senescence and aneurysm formation in MFS patients. This finding could serve as the basis for a novel strategy for treating aortic aneurysm in MFS.