Publications by authors named "Pei-Lin Shao"

51 Publications

Microplasma Treatment versus Negative Pressure Therapy for Promoting Wound Healing in Diabetic Mice.

Int J Mol Sci 2021 Sep 24;22(19). Epub 2021 Sep 24.

Department of Dermatology, National Cheng Kung University Hospital, College of Medicine, National Cheng Kung University, Tainan 704, Taiwan.

The delayed healing response of diabetic wounds is a major challenge for treatment. Negative pressure wound therapy (NPWT) has been widely used to treat chronic wounds. However, it usually requires a long treatment time and results in directional growth of wound healing skin tissue. We investigated whether nonthermal microplasma (MP) treatment can promote the healing of skin wounds in diabetic mice. Splint excision wounds were created on diabetic mice, and various wound healing parameters were compared among MP treatment, NPWT, and control groups. Quantitative analysis of the re-epithelialization percentage by detecting Ki67 and DSG1 expression in the extending epidermal tongue (EET) of the wound area and the epidermal proliferation index (EPI) was subsequently performed. Both treatments promoted wound healing by enhancing wound closure kinetics and wound bed blood flow; this was confirmed through histological analysis and optical coherence tomography. Both treatments also increased Ki67 and DSG1 expression in the EET of the wound area and the EPI to enhance re-epithelialization. Increased Smad2/3/4 mRNA expression was observed in the epidermis layer of wounds, particularly after MP treatment. The results suggest that the Smad-dependent transforming growth factor β signaling contributes to the enhancement of re-epithelialization after MP treatment with an appropriate exposure time. Overall, a short-term MP treatment (applied for 30 s twice a day) demonstrated comparable or better efficacy to conventional NPWT (applied for 4 h once a day) in promoting wound healing in diabetic mice. Thus, MP treatment exhibits promise for treating diabetic wounds clinically.
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http://dx.doi.org/10.3390/ijms221910266DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8508803PMC
September 2021

Additional benefit of induced pluripotent stem cell-derived mesenchymal stem cell therapy on sepsis syndrome-associated acute kidney injury in rat treated with antibiotic.

Stem Cell Res Ther 2021 10 7;12(1):526. Epub 2021 Oct 7.

Division of Cardiology, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University College of Medicine, 123, Dapi Road, Niaosung Dist., Kaohsiung City, 83301, Taiwan.

Background: This study tested whether human induced-pluripotent stem-cell-derived mesenchymal-stem-cells (iPS-MSCs) would offer an additional benefit to the rodent with acute kidney injury (AKI) (ischemia for 1 h followed by reperfusion for 120 h) associated sepsis syndrome (SS) (by cecal-ligation-puncture immediately after AKI-induction) undergoing ciprofloxacin therapy.

Results: Male-adult SD rats (n = 80) were categorized into group 1 (sham-operated-control, n = 10), group 2 (AKI + SS, n = 24), group 3 (AKI + SS + ciprofloxacin/3 mg/kg, orally for 120 h, n = 12), group 4 (AKI + SS + iPS-MSCs/1.2 × 10/intravenously administered by 3 h after AKI, n = 12), group 5 (AKI + SS + iPS-MSCs/1.2 × 10/intravenously administered by 18 h after AKI, n = 12), group 6 (AKI + SS + iPS-MSCs/1.2 × 10/intravenously administered by 3 h after AKI induction + ciprofloxacin, n = 10] and euthanized by 120 h. The result showed that the mortality was significantly higher in group 2 than in other groups (all p < 0.01). The creatinine level was highest in group 2, lowest in group 1, significantly lower in group 6 than in groups 3, 4 and 5, (all p < 0.0001), but it showed no difference among the latter 3 groups. Flow cytometric analysis showed that the circulatory inflammatory cells (Ly6G/CD11), early (AN-V+/PI-)/late (AN-V+/PI+) apoptosis, and circulatory/splenic immune cells (CD3+/CD4+, CD3+/CD8a+) were highest in group 2, lowest in group 1, significantly lower in group 6 than in groups 3/4/5 and significantly lower in group 4 than in groups 3/5 (all p < 0.0001), but they showed no difference between groups 3/5. Protein expressions of oxidative-stress (NOX-1/NOX2/oxidized protein), apoptotic (cleaved-caspase3/cleaved-PARP/mitochondrial-Bax), fibrotic (TGF-ß/Smad3), inflammatory (MMP-9/IL-6/TNF-α) and autophagic (Atg5/Beclin) biomarkers in kidney exhibited an identical pattern of circulatory inflammatory cells (all p < 0.0001).

Conclusion: Combined iPS-MSCs-ciprofloxacin therapy was superior to either one alone for protecting AKI complicated by SS.
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http://dx.doi.org/10.1186/s13287-021-02582-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8499569PMC
October 2021

Double overexpression of miR-19a and miR-20a in induced pluripotent stem cell-derived mesenchymal stem cells effectively preserves the left ventricular function in dilated cardiomyopathic rat.

Stem Cell Res Ther 2021 06 29;12(1):371. Epub 2021 Jun 29.

Division of Cardiology, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University, College of Medicine, 123, Dapi Road, Niaosung Dist, Kaohsiung, 83301, Taiwan.

Background: This study tested the hypothesis that double overexpression of miR-19a and miR-20a (dOex-mIRs) in human induced pluripotent stem cell (iPS)-derived mesenchymal stem cells (MSCs) effectively preserved left ventricular ejection fraction (LVEF) in dilated cardiomyopathy (DCM) (i.e., induced by doxorubicin) rat.

Methods And Results: In vitro study was categorized into groups G1 (iPS-MSC), G2 (iPS-MSC), G3 (iPS-MSC + HO/100uM), and G4 (iPS-MSC + HO/100uM). The in vitro results showed the cell viability was significantly lower in G3 than in G1 and G2, and that was reversed in G4 but it showed no difference between G1/G2 at time points of 6 h/24 h/48 h, whereas the flow cytometry of intra-cellular/mitochondrial oxidative stress (DCFA/mitoSOX) and protein expressions of mitochondrial-damaged (cytosolic-cytochrome-C/DRP1/Cyclophilin-D), oxidative-stress (NOX-1/NOX2), apoptotic (cleaved-caspase-3/PARP), fibrotic (p-Smad3/TGF-ß), and autophagic (ratio of LC3B-II/LC3BI) biomarkers exhibited an opposite pattern of cell-proliferation rate (all p< 0.001). Adult-male SD rats (n=32) were equally divided into groups 1 (sham-operated control), 2 (DCM), 3 (DCM + iPS-MSCs/1.2 × 10 cells/administered by post-28 day's DCM induction), and 4 (DCM + iPS-MSC/1.2 × 10 cells/administered by post-28 day's DCM induction) and euthanized by day 60 after DCM induction. LV myocardium protein expressions of oxidative-stress signaling (p22-phox/NOX-1/NOX-2/ASK1/p-MMK4,7/p-JNK1,2/p-cJUN), upstream (TLR-4/MAL/MyD88/TRIF/TRAM/ TFRA6/IKK/NF-κB) and downstream (TNF-α/IL-1ß/MMP-9) inflammatory signalings, apoptotic (cleaved-PARP/mitochondrial-Bax), fibrotic (Smad3/TGF-ß), mitochondrial-damaged (cytosolic-cytochrome-C/DRP1/cyclophilin-D), and autophagic (beclin1/Atg5) biomarkers were highest in group 2, lowest in group 1 and significantly lower in group 4 than in group 3, whereas the LVEF exhibited an opposite pattern of oxidative stress (all p< 0.0001).

Conclusion: iPS-MSC therapy was superior to iPS-MSC therapy for preserving LV function in DCM rat.
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http://dx.doi.org/10.1186/s13287-021-02440-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8243466PMC
June 2021

Uremic toxic substances are essential elements for enhancing carotid artery stenosis after balloon-induced endothelial denudation: worsening role of the adventitial layer.

Am J Transl Res 2020 15;12(11):7144-7159. Epub 2020 Nov 15.

Department of Nursing, Asia University Taichung 41354, Taiwan.

This study tested the hypothesis that uremic-toxic substances play a crucial role in enhancing left-common carotid artery (LCCA) stenosis after balloon-denudation of LCCA endothelium (BDLCCAE), and that the adventitial layer plays a complementary role in worsening LCCA stenosis. results showed the protein expressions of inflammation (IL-1β/TNF-α/IL-6), apoptosis (mitochondrial-Bax/cleaved-caspase-3/cleaved-PARP) and autophagy (beclin/Atg5/LC3B-II to LC3B-I ratio) as well as protein (NOX-1/NOX-2/p22phox/oxidized-protein), total cellular (H2DCFDA) and mitochondrial (Mitosox) levels of oxidative stress were significantly increased in p-Cresol-treated umbilical vein endothelial cells (HUVECs) as compared with control, whereas angiogenesis capacity (i.e., Matrigel-assay for HUVECs) exhibited an opposite pattern to inflammation between the two groups (all P < 0.001). Animals (n = 60) were categorized into group 1 (sham-operated control), group 2 (BDLCCAE), group 3 [BDLCCAE + ESRD patient's serum (1 cc/injection into deprived CA adventitia)], group 4 [BDLCCAE + ESRD patient's serum (1 cc/injection from peri-adventitia)], and group 5 [BDLCCAE + ESRD patient's serum (2 cc/by intravenous injection at days 1/3/7/10/14 after BDLCCADE)] and LCCA was harvested by day-21 after BDLCCAE procedure. Nitric-oxide release from LCCA and the LCCA cross-section area significantly and progressively reduced, whereas intimal and medial layers of LCCA significantly and progressively increased from groups 1 to 5 (all P < 0.001). The cellular expressions of inflammation (CD14+) and DNA-damage biomarker (γ-H2AX+) were significantly and progressively increased, whereas endothelial surface markers (CXCR4/vWF+) were significantly and progressively reduced from groups 1 to 5 (all P < 0.0001). Uremic toxins played an essential role in LCCA remodeling and obstruction. LCCA adventitia facilitated the initiation and propagation of LCCA proliferative obstruction.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7724323PMC
November 2020

Human Umbilical Cord-Derived Mesenchymal Stem Cell Therapy Effectively Protected the Brain Architecture and Neurological Function in Rat After Acute Traumatic Brain Injury.

Cell Transplant 2020 Jan-Dec;29:963689720929313

Department of Nursing, 63267Asia University, Taichung.

Intracranial hemorrhage from stroke and head trauma elicits a cascade of inflammatory and immune reactions detrimental to neurological integrity and function at cellular and molecular levels. This study tested the hypothesis that human umbilical cord-derived mesenchymal stem cell (HUCDMSC) therapy effectively protected the brain integrity and neurological function in rat after acute traumatic brain injury (TBI). Adult male Sprague-Dawley rats ( = 30) were equally divided into group 1 (sham-operated control), group 2 (TBI), and group 3 [TBI + HUCDMSC (1.2 × 10 cells/intravenous injection at 3 h after TBI)] and euthanized by day 28 after TBI procedure. The results of corner test and inclined plane test showed the neurological function was significantly progressively improved from days 3, 7, 14, and 28 in groups 1 and 3 than in group 2, and group 1 than in group 3 (all < 0.001). By day 28, brain magnetic resonance imaging brain ischemic volume was significantly increased in group 2 than in group 3 ( < 0.001). The protein expressions of apoptosis [mitochondrial-bax positive cells (Bax)/cleaved-caspase3/cleaved-poly(adenosine diphosphate (ADP)-ribose) polymerase], fibrosis (Smad3 positive cells (Smad3)/transforming growth factor-β), oxidative stress (NADPH Oxidase 1 (NOX-1)/NADPH Oxidase 2 (NOX-2)/oxidized-protein/cytochrome b-245 alpha chain (p22phox)), and brain-edema/deoxyribonucleic acid (DNA)-damaged biomarkers (Aquaporin-4/gamma H2A histone family member X ( (γ-H2AX)) displayed an identical pattern to neurological function among the three groups (all < 0.0001), whereas the protein expressions of angiogenesis biomarkers (vascular endothelial growth factor/stromal cell-derived factor-1α/C-X-C chemokine receptor type 4 (CXCR4)) significantly increased from groups 1 to 3 (all < 0.0001). The cellular expressions of inflammatory biomarkers (cluster of differentiation 14 (+) cells (CD14+)/glial fibrillary acidic protein positive cells (GFAP+)/ a member of a new family of EGF-TM7 molecules positive cells (F4/80+)) and DNA-damaged parameter (γ-H2AX) exhibited an identical pattern, whereas cellular expressions of neural integrity (hexaribonucleotide Binding Protein-3 positive cells (NeuN+)/nestin+/doublecortin+) exhibited an opposite pattern of neurological function among the three groups (all < 0.0001). Xenogeneic HUCDMSC therapy was safe and it significantly preserved neurological function and brain architecture in rat after TBI.
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http://dx.doi.org/10.1177/0963689720929313DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7784577PMC
August 2021

Dipeptidyl Peptidase-4 deficiency effectively protects the brain and neurological function in rodent after acute Hemorrhagic Stroke.

Int J Biol Sci 2020 16;16(16):3116-3132. Epub 2020 Oct 16.

Department of Anesthesiology, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University College of Medicine, Kaohsiung 83301, Taiwan.

This study tested the hypothesis that abrogated dipeptidyl peptidase-4 (DPP4) activity played a crucial role on reducing stroke volume and preserving neurological function in rodent after acute hemorrhagic stroke (AHS). Animals (n=6/each group) were categorized into group 1 (sham-control of F344 rat), group 2 (sham-control of DPP4-deficiency rat), group 3 [AHS by right cerebral injection of autologous blood (100 µL) in F344 rat], group 4 (AHS + sitagliptin/600 mg/kg 3 h prior to and at 3 h then once per day after AHS) and group 5 (AHS in DPP4-deficiency rat). The results of corner test showed the neurological function was significantly improved from days 3, 7, and 14 in groups 4 and 5 than in group 3 (all <0.001). By days 1 and 14 after AHS procedure, the circulating levels of SDF-1α and GLP-1 were significantly increased from groups 1/2 to group 5 (all <0.001), whereas circulating DPP4 activity was significantly increased in group 3 than other groups (all <0.001). The brain ischemic area (BIA) was highest in group 3, lowest in groups 1/2 and significantly lower in group 5 than in group 4 (all <0.0001). The protein expressions of oxidative-stress/inflammatory/apoptotic/cell-proliferation signaling, and the cellular expressions of inflammatory/DNA-damaged biomarkers exhibited a similar pattern to BIA among the groups (all <0.01). In conclusion, deprivation of DPP4 activity protected the brain from AHS damage and preserved neurological function.
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http://dx.doi.org/10.7150/ijbs.42677DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7645992PMC
November 2021

Circulatory Rejuvenated EPCs Derived from PAOD Patients Treated by CD34 Cells and Hyperbaric Oxygen Therapy Salvaged the Nude Mouse Limb against Critical Ischemia.

Int J Mol Sci 2020 Oct 23;21(21). Epub 2020 Oct 23.

Institute for Translational Research in Biomedicine, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung 83301, Taiwan.

This study tested whether circulatory endothelial progenitor cells (EPCs) derived from peripheral arterial occlusive disease (PAOD) patients after receiving combined autologous CD34+ cell and hyperbaric oxygen (HBO) therapy (defined as rejuvenated EPCs) would salvage nude mouse limbs against critical limb ischemia (CLI). Adult-male nude mice ( = 40) were equally categorized into group 1 (sham-operated control), group 2 (CLI), group 3 (CLI-EPCs (6 × 10) derived from PAOD patient's circulatory blood prior to CD34 cell and HBO treatment (EPC) by intramuscular injection at 3 h after CLI induction) and group 4 (CLI-EPCs (6 × 10) derived from PAOD patient's circulatory blood after CD34 cell and HBO treatment (EPC) by the identical injection method). By 2, 7 and 14 days after the CLI procedure, the ischemic to normal blood flow (INBF) ratio was highest in group 1, lowest in group 2 and significantly lower in group 4 than in group 3 ( < 0.0001). The protein levels of endothelial functional integrity (CD31/von Willebrand factor (vWF)/endothelial nitric-oxide synthase (eNOS)) expressed a similar pattern to that of INBF. In contrast, apoptotic/mitochondrial-damaged (mitochondrial-Bax/caspase-3/PARP/cytosolic-cytochrome-C) biomarkers and fibrosis (Smad3/TGF-ß) exhibited an opposite pattern, whereas the protein expressions of anti-fibrosis (Smad1/5 and BMP-2) and mitochondrial integrity (mitochondrial-cytochrome-C) showed an identical pattern of INBF (all < 0.0001). The protein expressions of angiogenesis biomarkers (VEGF/SDF-1α/HIF-1α) were progressively increased from groups 1 to 3 (all < 0.0010). The number of small vessels and endothelial cell surface markers (CD31/vWF) in the CLI area displayed an identical pattern of INBF (all < 0.0001). CLI automatic amputation was higher in group 2 than in other groups (all < 0.001). In conclusion, EPCs from HBO-C34+ cell therapy significantly restored the blood flow and salvaged the CLI in nude mice.
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http://dx.doi.org/10.3390/ijms21217887DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7660611PMC
October 2020

Early intramyocardial implantation of exogenous mitochondria effectively preserved left ventricular function in doxorubicin-induced dilated cardiomyopathy rat.

Am J Transl Res 2020 15;12(8):4612-4627. Epub 2020 Aug 15.

Department of Orthopedics, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University College of Medicine Kaohsiung 83301, Taiwan.

This study tested the hypothesis that early implantation of mitochondria (Mito) into left myocardium could effectively protect heart against doxorubicin/12 mg/kg-induced dilated cardiomyopathy (DCM) in rat. Adult-male SD rats (n = 18) were equally categorized into group 1 (sham control), group 2 (DCM) and group 3 [DCM + Mito (500 μg/rat intramyocardial injection by day-21 after DCM induction)] and euthanized by day 60. In vitro studies showed that exogenously-transferred Mito was abundantly identified in H9C2 cells. The q-PCR showed significant increase in relative number of mitDNA in Mito-transferred H9C2 cells than in control group (P<0.001). The mRNA-gene and protein expressions of NRF1/NRF2/Tfam/PGC-1α/ERRα/Mfn2 were significantly increased in low-dose Mito-transferred and more significantly increased in high-dose Mito-transferred H29C2 cells than in control group (all P<0.01). Day-60 left-ventricular-ejection-fraction (LVEF) was significantly lower in group 2 than in groups 1 and 3, and significantly lower in group 3 than in group 1 (P<0.0001). The ratios of lung and heart weights to tibial length and myocardial histopathological findings of fibrotic area/myocardial injured score/γ-H2AX+ cells exhibited an opposite pattern to LVEF among the three groups (all P<0.0001). The myocardial protein expressions of oxidative-stress (NOX-1/NOX-2/oxidized protein/p22phox), autophagic (beclin-1/Atg-5/ratio of CL3B-II/CL3B-I), and apoptotic/mitochondrial-damaged (cleaved-caspase-3/mitochondrial Bax/cleaved-PARP/cytosolic-cytochrome-C/DRP1/cyclophilin D1) biomarkers exhibited an opposite pattern, whereas the protein expressions of mitochondrial integrity (mitochondrial-cytochrome-C/mitochondrial-complex I/II/III/IV and Mfn2/PGC-1) exhibited an identical pattern to LVEF among the groups (all P<0.001). In conclusion, early Mito therapy effectively preserved LVEF and myocardial integrity in DCM rat.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7476136PMC
August 2020

Losing Regulation of the Extracellular Matrix is Strongly Predictive of Unfavorable Prognostic Outcome after Acute Myocardial Infarction.

Int J Mol Sci 2020 Aug 27;21(17). Epub 2020 Aug 27.

Division of Cardiology, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital, Chang Gung University College of Medicine, Kaohsiung 83301, Taiwan.

This study tested the hypothesis that MMP-9tPA double knock out (i.e., MT) plays a crucial role in the prognostic outcome after acute myocardial infarction (AMI by ligation of left-coronary-artery) in MT mouse. Animals were categorized into sham-operated controls in MT animals (group 1) and in wild type (B6: group 2), AMI-MT (group 3) and AMI-B6 (group 4) animals. They were euthanized, and the ischemic myocardium was harvested, by day 60 post AMI. The mortality rate was significantly higher in group 3 than in other groups and significantly higher in group 4 than in groups 1/2, but it showed no difference in the latter two groups (all < 0.01). By day 28, the left-ventricular (LV) ejection fraction displayed an opposite pattern, whereas by day 60, the gross anatomic infarct size displayed an identical pattern of mortality among the four groups (all < 0.001). The ratio of heart weight to tibial length and the lung injury score exhibited an identical pattern of mortality ( < 0.01). The protein expressions of apoptosis (mitochondrial-Bax/cleaved-caspase3/cleaved-PARP), fibrosis (Smad3/T-GF-ß), oxidative stress (NOX-1/NOX-2/oxidized-protein), inflammation (MMPs/TNF-α/p-NF-κB), heart failure/pressure overload (BNP/ß-MHC) and mitochondrial/DNA damage (cytosolic-cytochrome-C/γ-H2AX) biomarkers displayed identical patterns, whereas the angiogenesis markers (small vessel number/CD31+cells in LV myocardium) displayed opposite patterns of mortality among the groups (all < 0.0001). The microscopic findings of fibrotic/collagen deposition/infarct areas and inflammatory cell infiltration of LV myocardium were similar to the mortality among the four groups (all < 0.0001). MT strongly predicted unfavorable prognostic outcome after AMI.
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http://dx.doi.org/10.3390/ijms21176219DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7503422PMC
August 2020

Protective effect of combined therapy with hyperbaric oxygen and autologous adipose-derived mesenchymal stem cells on renal function in rodent after acute ischemia-reperfusion injury.

Am J Transl Res 2020 15;12(7):3272-3287. Epub 2020 Jul 15.

Division of Cardiology, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University College of Medicine Kaohsiung 83301, Taiwan.

Background: This study tested the hypothesis that combined hyperbaric oxygen (HBO) and autologous adipose-derived mesenchymal stem cell (ADMSC) therapy was superior to either alone at protecting renal function in rodents after acute ischemia-reperfusion (IR) injury.

Methods And Results: Adult-male SD rats (n = 40) were equally categorized: group 1 (sham-operated control); group 2 (IR + 50 μg medium intra-renal artery administration); group 3 [IR + HBO (at 1.5 h and days 1 and 2 after IR)]; group 4 [IR + ADMSC (2.0×10 cells/5.0×10/per each renal artery and 1.0×10 by intravenous injection at 1.5 h after IR]; and group 5 (IR + HBO-ADMSC). By 72 hr after IR, the circulating levels of BUN/creatinine and ratio of urine protein/creatinine were significantly highest in group 2, lowest in group 1, significantly increased in group 5 than in groups 3 and 4, but not different between latter two groups, whereas the circulating levels of EPCs and soluble-angiogenesis biomarkers (SDF-1α/HIF-1α) exhibited an opposite pattern to BUN/creatinine among the five groups (all P<0.001). The kidney injury score, ROS (fluorescent intensity of HDCFDA dye in kidney), inflammation (F4/80+, CD14+ cells) and glomerular-tubular injury score (WT-1/KIM-1) displayed an identical pattern whereas the integrity of podocyte components exhibited an opposite pattern to BUN/creatinine among the five groups (all P<0.0001). The protein expressions of inflammatory (MMP-9/TNF-α/NF-κB/ICAM-1), oxidative-stress (NOX-1/NOx-2/oxidized protein) and apoptotic (mitochondrial-Bax/cleaved-caspase3/PARP) markers showed an identical pattern to BUN/creatinine (all P<0.001).

Conclusion: Combined ADMSC-HBO therapy was superior to either one alone at protecting the kidney from acute IR injury.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7407680PMC
July 2020

Baseline factors identified for the prediction of good responders in patients with end-stage diffuse coronary artery disease undergoing intracoronary CD34+ cell therapy.

Stem Cell Res Ther 2020 07 29;11(1):324. Epub 2020 Jul 29.

Division of Cardiology, Department of Internal Medicine, College of Medicine, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University, No. 123, Ta Pei Road, Niao Sung District, Kaohsiung, 83301, Taiwan.

Background: Treating patients with end-stage diffuse coronary artery disease (EnD-CAD) unsuitable for coronary intervention remains a clinical challenge. They usually express refractory angina and have a high risk of mortality. Although growing data have indicated cell therapy is an alternative solution to medical or invasive therapy, there are still lacking useful markers to predict whether heart function will improve in the EnD-CAD patients who underwent circulatory-derived CD34+ cell therapy. By utilizing the baseline variables and results from our previous phase I/II clinical trials, the aim of this study tried to elucidate the variables predictive of the "good response" to CD34+ cell therapy.

Methods: This retrospective study included 38 patients in phase I clinical trial (2011-2014), and 30 patients in phase II clinical trial (2013-2017). These patients were categorized into "good responders" and "non-responders" according to their 1-year improvement of LVEF ≥ 7.0% or < 7.0% after intracoronary CD34+ cell therapy. Univariate and multivariate logistic regression models were performed to identify potential independent predictors of a good responder to cell therapy, followed by Hosmer-Lemeshow (H-L) test for goodness of fit and prediction power.

Results: Among baseline data, multivariate analysis demonstrated that the history of a former smoker was independently predictive of good responders (p = 0.006). On the other hand, male gender, the baseline Canadian Cardiovascular Society angina score ≥ 3, and grades of LV diastolic dysfunction ≥ 2 were significantly negative predictors of good responders (all p < 0.01). After administration of subcutaneous granulocyte-colony stimulating factor (G-CSF), a higher post-G-CSF neutrophil count in addition to the above four baseline variables also played crucial roles in early prediction of good response to CD34+ cell therapy for EnD-CAD (all p < 0.03). The H-L test displayed a good prediction power with sensitivity 83.3%, specificity 85.3%, and accuracy 84.4%.

Conclusions: Using the results of our phase I/II clinical trials, previous smoking habit, female sex, lower grades of angina score, and diastolic dysfunction were identified to be independently predictive of "good response" to CD34+ cell therapy in the patients with EnD-CAD.

Trial Registration: This is a retrospective analysis based on phase I ( ISRCTN72853206 ) and II ( ISRCTN26002902 ) clinical trials.
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http://dx.doi.org/10.1186/s13287-020-01835-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7391819PMC
July 2020

Intra-carotid arterial transfusion of circulatory-derived autologous endothelial progenitor cells in rodent after ischemic stroke-evaluating the impact of therapeutic time points on prognostic outcomes.

Stem Cell Res Ther 2020 06 5;11(1):219. Epub 2020 Jun 5.

Division of Cardiology, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University, College of Medicine, Kaohsiung, Taiwan.

Background: This study tested the optimal time point for left intra-carotid arterial (LICA) administration of circulatory-derived autologous endothelial progenitor cells (EPCs) for improving the outcome in rat after acute ischemic stroke (IS).

Methods And Results: Adult male SD rats (n = 70) were equally categorized into group 1 (sham-operated control), group 2 (IS), group 3 (IS+EPCs/1.2 × 10 cells/by LICA administration 3 h after IS), group 4 (IS+EPCs/LICA administration post-day-3 IS), group 5 (IS+EPCs/LICA administration post-day-7 IS), group 6 (IS+EPCs/LICA administration post-day-14 IS), and group 7 (IS+EPCs/LICA administration post-day-28 IS). The brain infarct volume (BIV) (at day 60/MRI) was lowest in group 1, highest in group 2, and significantly progressively increased from groups 3 to 7, whereas among the IS animals, the neurological function was significantly preserved in groups 3 to 6 than in groups 2 and 7 post-day-60 IS (all P < 0.0001). By day 60, the endothelial cell markers at protein and cellular levels and number of small vessels exhibited an opposite pattern of BIV among the groups (all P < 0.0001). The protein and cellular levels of inflammation, and protein levels of oxidative stress, autophagy, and apoptosis were highest in group 2, lowest in group 1, and progressively increased from groups 3 to 7 (all P < 0.0001). The angiogenesis biomarkers at protein and cellular levels were significantly progressively increased from groups 1 to 3, then significantly progressively decreased from groups 4 to 7 (all P < 0.0001).

Conclusion: Early EPC administration provided better benefits on improving functional/image/molecular-cellular outcomes after acute IS in rat.
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http://dx.doi.org/10.1186/s13287-020-01739-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7275327PMC
June 2020

Intravenous administration of iPS-MSC mobilized into CKD parenchyma and effectively preserved residual renal function in CKD rat.

J Cell Mol Med 2020 03 15;24(6):3593-3610. Epub 2020 Feb 15.

Institute for Translational Research in Biomedicine, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, Taiwan.

This study traced intravenously administered induced pluripotent stem cell (iPSC)-derived mesenchymal stem cells (MSC) and assessed the impact of iPSC-MSC on preserving renal function in SD rat after 5/6 nephrectomy. The results of in vitro study showed that FeraTrack™Direct contrast particles (ie intracellular magnetic labelling) in the iPSC-MSC (ie iPS-MSC ) were clearly identified by Prussian blue stain. Adult-male SD rats (n = 40) were categorized into group 1 (SC), group 2 [SC + iPS-MSC (1.0 × 10 cells)/intravenous administration post-day-14 CKD procedure], group 3 (CKD), group 4 [CKD + iPS-MSC (0.5 × 10 cells)] and group 5 [CKD + iPS-MSC (1.0 × 10 cells)]. By day-15 after CKD induction, abdominal MRI demonstrated that iPS-MSC were only in the CKD parenchyma of groups 4 and 5. By day 60, the creatinine level/ratio of urine protein to urine creatinine/kidney injury score (by haematoxylin and eosin stain)/fibrotic area (Masson's trichrome stain)/IF microscopic finding of kidney injury molecule-1 expression was lowest in groups 1 and 2, highest in group 3, and significantly higher in group 4 than in group 5, whereas IF microscopic findings of podocyte components (ZO-1/synaptopodin) and protein levels of anti-apoptosis ((Bad/Bcl-xL/Bcl-2) exhibited an opposite pattern to creatinine level among the five groups (all P < .0001). The protein expressions of cell-proliferation signals (PI3K/p-Akt/m-TOR, p-ERK1/2, FOXO1/GSK3β/p90RSK), apoptotic/DNA-damage (Bax/caspases8-10/cytosolic-mitochondria) and inflammatory (TNF-α/TNFR1/TRAF2/NF-κB) biomarkers displayed an identical pattern to creatinine level among the five groups (all P < .0001). The iPS-MSC that were identified only in CKD parenchyma effectively protected the kidney against CKD injury.
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http://dx.doi.org/10.1111/jcmm.15050DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7131913PMC
March 2020

Soluble ST2 is a Useful Biomarker for Grading Cerebral-Cardiac Syndrome in Patients after Acute Ischemic Stroke.

J Clin Med 2020 Feb 11;9(2). Epub 2020 Feb 11.

Division of Cardiology, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University, College of Medicine, Kaohsiung 83301, Taiwan.

This study tested whether the soluble (s)ST2 is a superb biomarker predictive of moderate to severe cerebral-cardiac syndrome (CCS) (defined as coexisting National Institute of Health Stroke Scale (NIHSS) >8 and left-ventricular ejection fraction (LVEF) <60%) in patients after acute ischemic stroke (IS). Between November 2015 and October 2017, a total of 99 IS patients were prospectively enrolled and categorized into three groups based on NIHSS, i.e., group 1 (NIHSS ≤ 8, = 66), group 2 (NIHSS = 9-15, = 14) and group 3 (NIHSS ≥ 16, = 19), respectively. Blood samples were collected immediately after hospitalization, followed by transthoracic echocardiographic examination. The results showed that the flow cytometric analysis for assessment of inflammatory biomarkers of TLR2+/CD14+cells, TLR4+/CD14+cells, Ly6g+/CD14+cells, and MPO+/CD14+cells, and ELISA assessment for circulatory level of sST2 were significantly higher in groups 2/3 than in group 1 (all < 0.01). However, these parameters did not show significant differences between groups 2 and 3 (all > 0.05). The LVEF was significantly lower in group 3 than in group 1 ( < 0.001), but it displayed no difference between groups 1/2 or between groups 2/3. These inflammatory biomarkers ((TLR2+/CD14+cells// TLR4+/CD14+cells// MPO+/CD14+cells) and sST2)) were significantly positively correlated to NIHSS and strongly negatively correlated to LVEF (all < 0.05). Multivariate analysis demonstrated that both MPO/CD14+cells >20% ( = 0.027) and sST2 ≥ 17,600 ( = 0.004) were significantly and independently predictive of moderate-severe CCS after acute IS. Receiver operating characteristic curve analysis demonstrated that sST2 was the most powerful predictor of CCS with a sensitivity of 0.929 and a specificity of 0.731 ( < 0.001). In conclusion, sST2 is a useful biomarker for prediction of CCS severity in patients after acute IS.
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http://dx.doi.org/10.3390/jcm9020489DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7074380PMC
February 2020

Human induced pluripotent stem cell-derived mesenchymal stem cell therapy effectively reduced brain infarct volume and preserved neurological function in rat after acute intracranial hemorrhage.

Am J Transl Res 2019 15;11(9):6232-6248. Epub 2019 Sep 15.

Division of Cardiology, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University College of Medicine Kaohsiung 83301, Taiwan.

We tested the hypothesis that human induced pluripotent stem cell-derived mesenchymal stem cell (iPSC-MSC) therapy could effectively reduce brain-infarct volume (BIV) and improve neurological function in rat after acute intracranial hemorrhage (ICH) induced by a weight-drop device. Adult-male SD rats (n=40) were equally divided into group 1 (sham-operated control), group 2 (ICH), group 3 (ICH + hyaluronic acid (HA)/intracranial injection at 3 h after ICH), group 4 [ICH + HA + iPSC-MSC (1.2 × 10 cells/intracranial injection at 3 h after ICH)] and euthanized by day 28 after ICH procedure. In vitro study showed that hemorrhagic-brain tissue augmented protein expressions of inflammation (HMGB1/MyD88/TLR-4/TLR-2/NF-κB/TNF-α/iNOS/IL-1β) in cultured neurons that were significantly inhibited by iPSC-MSC treatment (all P<0.001). By days 7 and 14 after ICH procedure, circulating inflammatory levels of TNF-α/IL-6/MPO expressed were lowest in group 1, highest in group 2 and significantly lower in group 4 than in group 3 (all P<0.0001). By day 14 after ICH procedure, neurological function and BIV expressed an opposite pattern, whereas protein expressions of inflammation (HMGB1/MyD88/TLR-4/TLR-2/NF-κB/I-kB/TNF-α/iNOS/IL-1β/MMP-9), oxidative stress (NOX-1/NOX-2/oxidized protein) and apoptosis (mitochondrial-Bax/cleaved-caspase-2/PARP) in brain exhibited an identical pattern to circulating inflammation among the four groups (all P<0.001). Microscopy demonstrated that the number of vascular remodeling and GFAP+/53BP1+/γ-H2AX+ cells displayed an identical pattern of inflammation, whereas the NeuN+ cells displayed an opposite pattern of inflammation among the four groups (all P<0.001). In conclusion, iPSC-MSC therapy markedly reduced BIV and preserved neurological function mainly by inhibiting inflammatory/oxidative-stress generation.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6789217PMC
September 2019

Early administration of cold water and adipose derived mesenchymal stem cell derived exosome effectively protects the heart from ischemia-reperfusion injury.

Am J Transl Res 2019 15;11(9):5375-5389. Epub 2019 Sep 15.

Division of Cardiology, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University College of Medicine Kaohsiung 83301, Taiwan.

This study tested the hypothesis that early administration with cold water (CW)-assisted adipose-derived mesenchymal stem cell (ADMSC)-derived exosome (Exo) therapy was superior to either one on protecting the heart against ischemia-reperfusion (IR) (i.e., by ligation of 50 minutes and relieved by day 5 prior to euthanizing the animals) injury. Adult-male SD rats (n=30) were equally categorized into groups 1 (sham-operated control), 2 (IR), 3 (IR + CW), 4 (IR + Exo) and 5 (IR + CW-Exo). The left ventricular ejection fraction (LVEF) was highest in group 1, lowest in group 2, and significantly higher in group 5 than in groups 3 and 4, but no difference between groups 3 and 4 (all P<0.001). The protein expressions of oxidative-stress (NOX-1/NOX-2/NOX-4/oxidized protein), apoptotic/mitochondrial-damaged (mitochondrial-Bax/caspase 3/PARP/p53/cytosolic-cytochrome-C) and inflammatory (IL-1β/TNF-α/NF-κB/MMP-9) biomarkers, and cellular-stress response signaling (PI3K/Akt/GSK3β and p-m-TOR) showed an opposite pattern, whereas the anti-oxidants (SIRT1/SIRT3), anti-inflammation (IL-10) and IKB-α/p-AMKP/mitochondrial-cytochrome-C exhibited an identical pattern to the LVEF among the five groups (all P<0.0001). The cellular expressions of inflammation (CD68), total cellular ROS (i.e., stained by HDCFDA) and the LV infarct/fibrotic/collagen-deposition areas displayed an opposite pattern, whereas the cell gap junction (coonexin 43) and sarcomere length exhibited an identical pattern of LVEF among the five groups (all P<0.0001). Conclusion: Combined CW-exosome therapy markedly protected the heart against IR injury.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6789220PMC
September 2019

Preactivated and disaggregated shape-changed platelets protect kidney against from ischemia-reperfusion injury in rat through attenuating inflammation reaction.

J Tissue Eng Regen Med 2019 12 22;13(12):2155-2168. Epub 2019 Oct 22.

Center for Shockwave Medicine and Tissue Engineering, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, Taiwan.

This study tested the hypothesis that preactivated and disaggregated shape-changed platelet (PreD-SCP) therapy significantly protected rat kidney from ischemia-reperfusion (IR) injury. Adult-male Sprague-Dawley rats (n = 24) were equally categorized into Groups 1 (sham-operated control [SC]), 2 (SC + PreD-SCP), 3 (IR only), and 4 (IR + PreD-SCP). By 72 hr after IR procedure, the circulatory levels of creatinine, blood urine nitrogen and inflammatory biomarkers (interleukin [IL]-6/tumor necrosis factor [TNF]-α), and ratio of urine protein to urine creatinine were significantly higher in Group 3 than in other groups and significantly higher in Group 4 than in Groups 1 and 2, but they showed no different between Groups 1 and 2 (all p < .001). The microscopic findings showed that the expressions of kidney injury score, cellular inflammation (MMP-9/CD14//F4/80), and fibrotic area were identical to the circulatory inflammation, whereas the integrity of podocyte components (ZO-1/synaptopodin/podocin) exhibited an opposite to circulatory inflammation among the four groups (all p < .0001). The protein expressions of inflammatory (TNF-α/IL-1ß/NF-κB/iNOS/TRAF6/MyD88/TLR-4), apoptotic/cell death (mitochondrial Bax/cleaved caspase-3/p-53), oxidized protein, mitogen-activated protein kinase family (p-38/p-JNK/p-c-JUN), and mitochondrial-damaged biomarkers displayed a similar pattern, whereas the antiapoptotic (Bcl-2/Bcl-XL) and integrity of mitochondrial biomarkers followed an opposite trend to circulatory inflammation among the four groups (all p < .001). PreD-SCP therapy effectively protected the kidney against IR injury.
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http://dx.doi.org/10.1002/term.2960DOI Listing
December 2019

The therapeutic impact of entresto on protecting against cardiorenal syndrome-associated renal damage in rats on high protein diet.

Biomed Pharmacother 2019 Aug 17;116:108954. Epub 2019 May 17.

Center for Shockwave Medicine and Tissue Engineering, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, 83301, Taiwan, ROC; Division of Cardiology, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital, Chang Gung University College of Medicine, Kaohsiung, 83301, Taiwan, ROC; Department of Nursing, Asia University, Taichung, 41354, Taiwan, ROC; Department of Medical Research, China Medical University Hospital, China Medical University, Taichung, 40402, Taiwan, ROC; Institute for Translational Research in Biomedicine, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, 83301, Taiwan, ROC. Electronic address:

Background: This study tested the hypothesis that Entresto could safely and effectively preserve heart and kidney function in rats with cardiorenal syndrome (CRS) induced by 5/6 nephrectomy and intra-peritoneal doxorubicin administration (accumulated dosage up to 7.5 mg/kg) together with daily high-protein-diet (H).

Methods And Results: Adult male Sprague-Dawley rats (n = 24) were equally categorized into Group 1 (sham-operated control + H), Group 2 (CRS + H) and Group 3 [CRS + H + Entresto (100 mg/kg/day orally) since Day 14 after CRS induction] and euthanized by Day 63 after CRS induction. By Day 63, circulatory BUN and creatinine levels and ratios of urine protein to creatinine were significantly higher in Group 2 than those in Groups 1 and 3, and significantly higher in Group 3 than in Group 1, whereas left-ventricular ejection fraction and kidney weight showed an opposite pattern among all groups (all p < 0.001). Microscopically, fibrosis area and intensity of oxidative stress (i.e., DCFDA stain) in kidney/heart tissues exhibited a pattern identical to that of creatinine level among all groups (all p < 0.0001). Kidney injury score and protein expressions of autophagy (i.e., beclin-1/Atg-5/protein ratio of LC3-BII/LC3-BI), fibrosis (Smad3/TGF-ß), apoptosis (mitochondrial-Bax/capase2/3/9), oxidative-stress (NOX-4/oxidized protein/xanthine-oxidase/catalase), membranous p47phox phosphorylation and mitochondrial-damage biomarker (cytosolic-cytochrome-C) were higher in Group 2 than those in Groups 1 and 3, and significantly higher in Group 3 than in Group 1, while protein expressions of anti-apoptosis (Bcl-2/Bcl-XL) and mitochondrial integrity (mitochondrial-cytochrome-C) markers displayed an opposite pattern among all groups in kidney tissues (all p < 0.0001).

Conclusion: Oral administration of entresto was safe and could offer protection against CRS-induced heart and kidney damage.
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http://dx.doi.org/10.1016/j.biopha.2019.108954DOI Listing
August 2019

Hyperbaric oxygen facilitates the effect of endothelial progenitor cell therapy on improving outcome of rat critical limb ischemia.

Am J Transl Res 2019 15;11(4):1948-1964. Epub 2019 Apr 15.

Department of Orthopedics, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University College of Medicine Kaohsiung 83301, Taiwan.

We tested the hypothesis that hyperbaric oxygen (HBO) (100% oxygen/2.4 atmospheres) facilitated the effect of autologous endothelial progenitor cell (EPC) therapy on restoring the blood flow in rat critical-limb ischemia (CLI). Adult-male-SD rats (n = 8/each group) were categorized into group 1 [sham control (SC)], group 2 (CLI-treated with culture medium), group 3 [CLI-intermittent HBO (3 h/day for 5 consecutive days after CLI), group 4 (CLI-EPC/2.0 × 10 cells), and group 5 (CLI-HBO-EPC). By day 5 after CLI, flow cytometry showed that the circulating EPC (Sca-1/CD31+/C-kit/CD31+/CD34+) levels were highest in group 5 and lowest in group 2 (all < 0.001). By day 14, laser Doppler demonstrated that the ratio of blood flow (i.e., CLI to normal hind-limb) was highest in group 1, lowest in group 2 and significantly higher in group 5 than in groups 3 and 4 (all < 0.0001). The protein expressions of endothelial-cell biomarkers (CD31/vWF/eNOS), and numbers of endothelial-cell markers (CD31+/vWF+) and small vessels exhibited a similar pattern to blood-flow ratio among five groups, whereas the angiogenesis parameters in protein (CXCR4/SDF-1α/HIF-1α/VEGF) and cellular (HIF-1α/SDF-1α/CXCR4+) levels were progressively increased from groups 1 to 5 (all < 0.0001). The protein expression of apoptotic (mitochondrial-Bax/cleaved-capspase-3/PARP), fibrotic (p-Smad3/TGF-ß) and mitochondrial-damaged (cytosolic-cytochrome C) exhibited an opposite pattern, whereas the protein expressions of anti-fibrotic (BMP-2/p-Smad1/5) and mitochondrial integrity (mitochondrial-cytochrome C) exhibited an identical pattern of ratio of blood flow among the five groups (all < 0.0001). Combined HBO-EPC therapy is superior to either one alone in improving ischemia in rodent CLI.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6511789PMC
April 2019

Alpha-5 Integrin Mediates Simvastatin-Induced Osteogenesis of Bone Marrow Mesenchymal Stem Cells.

Int J Mol Sci 2019 Jan 24;20(3). Epub 2019 Jan 24.

Orthopaedic Research Center, College of Medicine, Kaohsiung Medical University, Kaohsiung 807, Taiwan.

Simvastatin (SVS) promotes the osteogenic differentiation of mesenchymal stem cells (MSCs) and has been studied for MSC-based bone regeneration. However, the mechanism underlying SVS-induced osteogenesis is not well understood. We hypothesize that α5 integrin mediates SVS-induced osteogenic differentiation. Bone marrow MSCs (BMSCs) derived from BALB/C mice, referred to as D1 cells, were used. Alizarin red S (calcium deposition) and alkaline phosphatase (ALP) staining were used to evaluate SVS-induced osteogenesis of D1 cells. The mRNA expression levels of α5 integrin and osteogenic marker genes (bone morphogenetic protein-2 (BMP-2), runt-related transcription factor 2 (Runx2), collagen type I, ALP and osteocalcin (OC)) were detected using quantitative real-time PCR. Surface-expressed α5 integrin was detected using flow cytometry analysis. Protein expression levels of α5 integrin and phosphorylated focal adhesion kinase (p-FAK), which is downstream of α5 integrin, were detected using Western blotting. siRNA was used to deplete the expression of α5 integrin in D1 cells. The results showed that SVS dose-dependently enhanced the gene expression levels of osteogenic marker genes as well as subsequent ALP activity and calcium deposition in D1 cells. Upregulated p-FAK was accompanied by an increased protein expression level of α5 integrin after SVS treatment. Surface-expressed α5 integrin was also upregulated after SVS treatment. Depletion of α5 integrin expression significantly suppressed SVS-induced osteogenic gene expression levels, ALP activity, and calcium deposition in D1 cells. These results identify a critical role of α5 integrin in SVS-induced osteogenic differentiation of BMSCs, which may suggest a therapeutic strategy to modulate α5 integrin/FAK signaling to promote MSC-based bone regeneration.
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http://dx.doi.org/10.3390/ijms20030506DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6387019PMC
January 2019

Shock Wave Therapy Enhances Mitochondrial Delivery into Target Cells and Protects against Acute Respiratory Distress Syndrome.

Mediators Inflamm 2018 21;2018:5425346. Epub 2018 Oct 21.

Center for Shockwave Medicine and Tissue Engineering, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung 83301, Taiwan.

This study tested the hypothesis that shock wave therapy (SW) enhances mitochondrial uptake into the lung epithelial and parenchymal cells to attenuate lung injury from acute respiratory distress syndrome (ARDS). ARDS was induced in rats through continuous inhalation of 100% oxygen for 48 h, while SW entailed application 0.15 mJ/mm for 200 impulses at 6 Hz per left/right lung field. In vitro and ex vivo studies showed that SW enhances mitochondrial uptake into lung epithelial and parenchyma cells (all < 0.001). Flow cytometry demonstrated that albumin levels and numbers of inflammatory cells (Ly6G+/CD14+/CD68+/CD11+) in bronchoalveolar lavage fluid were the highest in untreated ARDS, were progressively reduced across SW, Mito, and SW + Mito (all < 0.0001), and were the lowest in sham controls. The same profile was also seen for fibrosis/collagen deposition, levels of biomarkers of oxidative stress (NOX-1/NOX-2/oxidized protein), inflammation (MMP-9/TNF-/NF-B/IL-1/ICAM-1), apoptosis (cleaved caspase 3/PARP), fibrosis (Smad3/TGF-), mitochondrial damage (cytosolic cytochrome c) (all < 0.0001), and DNA damage (-H2AX+), and numbers of parenchymal inflammatory cells (CD11+/CD14+/CD40L+/F4/80+) ( < 0.0001). These results suggest that SW-assisted Mito therapy effectively protects the lung parenchyma from ARDS-induced injury.
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http://dx.doi.org/10.1155/2018/5425346DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6215567PMC
February 2019

Extracorporeal Shock Wave-Supported Adipose-Derived Fresh Stromal Vascular Fraction Preserved Left Ventricular (LV) Function and Inhibited LV Remodeling in Acute Myocardial Infarction in Rat.

Oxid Med Cell Longev 2018 17;2018:7518920. Epub 2018 Oct 17.

Division of Cardiology, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University College of Medicine, Kaohsiung 83301, Taiwan.

This study tested the hypothesis that extracorporeal shock wave- (ECSW-) assisted adipose-derived stromal vascular fraction (SVF) therapy could preserve left ventricular ejection fraction (LVEF) and inhibit LV remodeling in a rat after acute myocardial infarction (AMI). Adult male SD rats were categorized into group 1 (sham control), group 2 (AMI induced by left coronary artery ligation), group 3 [AMI + ECSW (280 impulses at 0.1 mJ/mm, applied to the chest wall at 3 h, days 3 and 7 after AMI), group 4 [AMI + SVF (1.2 × 10) implanted into the infarct area at 3 h after AMI], and group 5 (AMI + ECSW-SVF). In vitro, SVF protected H9C2 cells against menadione-induced mitochondrial damage and increased fluorescent intensity of mitochondria in nuclei ( < 0.01). By day 42 after AMI, LVEF was highest in group 1, lowest in group 2, significantly higher in group 5 than in groups 3 and 4, and similar between the latter two groups (all < 0.0001). LV remodeling and infarcted, fibrotic, and collagen deposition areas as well as apoptotic nuclei exhibited an opposite pattern to LVEF among the groups (all < 0.0001). Protein expressions of CD31/vWF/eNOS/PGC-1/-MHC/mitochondrial cytochrome C exhibited an identical pattern, whilst protein expressions of MMP-9/TNF-/IL-1/NF-B/caspase-3/PARP/Samd3/TGF-/NOX-1/NOX-2/oxidized protein/-MHC/BNP exhibited an opposite pattern to LVEF among five groups (all < 0.0001). Cellular expressions of CXCR4/SDF-1/Sca-1/c-Kit significantly and progressively increased from groups 1 to 5 (all < 0.0001). Cellular expression of -H2AX/CD68 displayed an opposite pattern to LVEF among the five groups (all < 0.0001). In conclusion, ECSW-SVF therapy effectively preserved LVEF and inhibited LV remodeling in rat AMI.
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http://dx.doi.org/10.1155/2018/7518920DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6207868PMC
December 2018

Melatonin-mediated downregulation of ZNF746 suppresses bladder tumorigenesis mainly through inhibiting the AKT-MMP-9 signaling pathway.

J Pineal Res 2019 Jan 18;66(1):e12536. Epub 2018 Nov 18.

Department of Nursing, Asia University, Taichung, Taiwan.

There still lacking effective treatment for bladder cancer. This study investigated whether melatonin (Mel) can suppress the growth and invasion of bladder cancer cells. Male C57B/L6 mice were categorized into control group (ie, subcutaneous injection of HT1197 bladder cancer cell line at the back] and treatment group [subcutaneous HT1197 cells + intraperitoneal Mel (100 mg/kg/d) from day 8 to day 21 after tumor cell injection]. In vitro Mel suppressed cell growth of four bladder cancer cell lines (ie, T24, RT4, HT1197, HT1376), cell migration in HT1197/HT1376, mitochondrial membrane potential (MMP) in T24 and colony formation in RT4 cells as well as arrested the cell cycle at G0 phase and inhibited the mitotic phase of T24 cells (all P < 0.0001). Protein expression of ZNF746 in RT4/T24 cells and protein expression phosphorylated (p)-AKT/MMP-2/MMP-9 in HT1197/HT1376 cells were reduced following Mel treatment (all P < 0.001). Transfection of T24 cells with plasmid-based shRNA (ie, ZNF746-silencing) downregulated the protein expression of MMP-9, cell growth, and invasion and attachment to endothelial cells but upregulated the colony formation (all P < 0.001). Mel suppressed oxidative stress and MMP but upregulated mitochondria mass in ZNF746-silenced T24 cells, whereas these parameters exhibited a similar patter to Mel treatment in ZNF746-silenced T24 cells (all P < 0.0001). In vivo study demonstrated that Mel treatment significantly suppressed cellular expressions of MMP-9/MMP-2, protein expressions of ZNF746/p-AKT, and tumor size (all P < 0.001). Mel treatment suppressed the growth, migration, and invasion of bladder carcinoma cells through downregulating ZNF746-regulated MMP-9/MMP-2 signaling.
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http://dx.doi.org/10.1111/jpi.12536DOI Listing
January 2019

Combined Therapy with SS31 and Mitochondria Mitigates Myocardial Ischemia-Reperfusion Injury in Rats.

Int J Mol Sci 2018 Sep 15;19(9). Epub 2018 Sep 15.

Division of Cardiovascular Surgery, Department of Surgery, Tri-Service General Hospital, National Defense Medical Center, Taipei 11490, Taiwan.

Myocardial ischemia-reperfusion (IR) injury contributes to adverse cardiac outcomes after myocardial ischemia, cardiac surgery, or circulatory arrest. In this study, we evaluated the ability of combined SS31-mitochondria (Mito) therapy to protect heart cells from myocardial IR injury. Adult male SD rats ( = 8/each group) were randomized: group 1 (sham-operated control), group 2 (IR, 30-min ischemia/72 h reperfusion), group 3 (IR-SS31 (2 mg intra-peritoneal injection at 30 min/24 h/48 h after IR)), group 4 (IR-mitochondria (2 mg/derived from donor liver/intra-venous administration/30 min after IR procedure)), and group 5 (IR-SS31-mitochondria). In H9C2 cells, SS31 suppressed menadione-induced oxidative-stress markers (NOX-1, NOX-2, oxidized protein) while it increased SIRT1/SIRT3 expression and ATP levels. In adult male rats 72 h after IR, left ventricular ejection fraction (LVEF) was highest in sham-operated control animals and lowest in the IR group. LVEF was also higher in IR rats treated with SS31-Mito than untreated IR rats or those treated with Mito or SS31 alone. Areas of fibrosis/collagen-deposition showed the opposite pattern. Likewise, levels of oxidative-stress markers (NOX-1, NOX-2, oxidized protein), inflammatory markers (MMP-9, CD11, IL-1β, TNF-α), apoptotic markers (mitochondrial-Bax, cleaved-caspase-3, PARP), fibrosis markers (p-Smad3, TGF-β), DNA-damage (γ-H2AX), sarcomere-length, and pressure/volume overload markers (BNP, β-MHC) all showed a pattern opposite that of LVEF. Conversely, anti-apoptotic (BMP-2, Smad1/5) and energy integrity (PGC-1α/mitochondrial cytochrome-C) markers exhibited a pattern identical to that of LVEF. This study demonstrates that the combined SS31-Mito therapy is superior to either therapy alone for protecting myocardium from IR injury and indicates that the responsible mechanisms involved increased SIRT1/SIRT3 expression, which suppresses inflammation and oxidative stress and protects mitochondrial integrity.
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http://dx.doi.org/10.3390/ijms19092782DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6164143PMC
September 2018

Combined Therapy with Extracorporeal Shock Wave and Adipose-Derived Mesenchymal Stem Cells Remarkably Improved Acute Ischemia-Reperfusion Injury of Quadriceps Muscle.

Oxid Med Cell Longev 2018 2;2018:6012636. Epub 2018 Apr 2.

Division of Cardiology, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University College of Medicine, Kaohsiung 83301, Taiwan.

Extracorporeal shock wave (ECSW) and adipose-derived mesenchymal stem cells (ADMSCs) have been recognized to have capacities of anti-inflammation and angiogenesis. We tested the hypothesis that ECSW and ADMSC therapy could attenuate ischemia-reperfusion- (IR-) induced thigh injury (femoral artery tightened for 6 h then the tightness was relieved) in rats. Adult male SD rats ( = 30) were divided into group 1 (sham-control), group 2 (IR), group 3 (IR + ECSW/120 impulses at 0.12 mJ/mm given at 3 h/24 h/72 h after IR), group 4 (allogenic ADMSC/1.2 × 10 cell intramuscular and 1.2 × 10 cell intravenous injections 3 h after IR procedure), and group 5 (ECSW + ADMSC). At day 7 after the IR procedure, the left quadriceps muscle was harvested for studies. At 18 h after the IR procedure, serum myoglobin/creatine phosphokinase (CPK) levels were highest in group 2, lowest in group 1, and with intermediate values significantly progressively reduced in groups 3 to 5 (all < 0.0001). By day 5 after IR, the mechanical paw-withdrawal threshold displayed an opposite pattern of CPK (all < 0.0001). The protein expressions of inflammatory, oxidative-stress, apoptotic, fibrotic, DNA-damaged, and mitochondrial-damaged biomarkers and cellular expressions of inflammatory and DNA-damaged biomarkers exhibited an identical pattern of CPK among the five groups (all < 0.0001). The microscopic findings of endothelial-cell biomarkers and number of arterioles expressed an opposite pattern of CPK, and the angiogenesis marker was significantly progressively increased from groups 1 to 5, whereas the histopathology showed that muscle-damaged/fibrosis/collagen-deposition areas exhibited an identical pattern of CPK among the five groups (all < 0.0001). In conclusion, ECSW-ADMSC therapy is superior to either one applied individually for protecting against IR-induced thigh injury.
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http://dx.doi.org/10.1155/2018/6012636DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5901825PMC
October 2018

Adipose-derived mesenchymal stem cell-derived exosomes alleviate overwhelming systemic inflammatory reaction and organ damage and improve outcome in rat sepsis syndrome.

Am J Transl Res 2018 15;10(4):1053-1070. Epub 2018 Apr 15.

Division of Colorectal Surgery, Department of Surgery, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University College of Medicine Kaohsiung 83301, Taiwan, China.

This study tested the hypothesis that healthy adipose-derived mesenchymal stem cell (ADMSC)-derived exosomes (HMSC) and apoptotic (A) (induced by 12 h hypoxia/12 h starvation)-ADMSC-derived exosomes (AMSC) were comparably effective at alleviating sepsis syndrome [SS; induced by cecal-ligation and puncture (CLP)]-induced systemic inflammation and reduced organ damage and unfavorable outcomes in rats. SD rats were divided into sham control (SC), SS only, SS + HMSC (100 µg intravenous administration 3 h after CLP), and AMSC. By day 5 after CLP procedure, the mortality rate was significantly higher in SS than in SC and HMSC (all P < 0.01), but it showed no significant different between SC and HMSC, between AMSC and HMSC or between SS and AMSC (P > 0.05). The levels of inflammatory mediators in circulation (CD11/Ly6G/MIF), bronchioalveolar lavage (CD11/Ly6G) and abdominal ascites (CD11/CD14/Ly6G/MIF) were highest in SS, lowest in SC and significantly higher in AMSC than in HMSC (all P < 0.001). The circulating/splenic levels of immune cells (CD34+/CD4+/CD3+/CD8+) were expressed in an identical pattern whereas the T-reg+ cells exhibited an opposite pattern of inflammation among the groups (all P < 0.001). The protein expressions of inflammation (MMP-9/MIF/TNF-α/NF-κB/IL-1β) and oxidative stress (NOX-1/NOX-2/oxidized protein), and cellular expressions (CD14+/CD68+) in lung/kidney parenchyma exhibited an identical pattern of inflammatory mediators (all P < 0.001). The kidney/lung injury scores displayed an identical pattern of inflammatory mediators among the groups (all P < 0.001). In conclusion, HMSC might be superior to AMSC for improving survival and suppressing the inflammatory reactions in rats after SS.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5934566PMC
April 2018

Extracorporeal shock wave markedly alleviates radiation-induced chronic cystitis in rat.

Am J Transl Res 2018 15;10(3):1036-1052. Epub 2018 Mar 15.

Divisions of General Medicine, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University College of MedicineKaohsiung 83301, Taiwan.

This study tested the hypothesis that extracorporeal shock wave (ECSW) treatment can effectively inhibit radiation-induced chronic cystitis (CC). Adult male Sprague-Dawley (SD) rats (n = 24) were randomly divided into group 1 (normal control), group 2 (CC induced by radiation with 300 cGy twice with a four-hour interval to the urinary bladder), group 3 [CC with ECSW treatment (0.2 mJ/mm/120 impulses/at days 1, 7, and 14 after radiation)]. Bladder specimens were harvested by day 28 after radiation. By day 28 after radiation, the degree of detrusor contraction impairment was significantly higher in group 2 than that in groups 1 and 3, and significantly higher in group 3 than that in group 1 (P<0.0001). The urine albumin concentration expressed an opposite pattern compared to that of detrusor function among the three groups (P<0.0001). The bladder protein expressions of inflammatory (TLR-2/TLR-4/IL-6/IL-12/MMP-9/TNF-α/NF-κB/RANTES/iNOS) and oxidative-stress (NOX-1/NOX-2/oxidized protein) biomarkers exhibited a pattern identical to that of urine albumin in all groups (all P<0.0001). The cellular expressions of inflammatory (CD14+/CD68+/CD74+/COX-2/MIF+/substance P+) and cytokeratin (CK13+/HMW CK+/CK+17/CK+18/CK+19) biomarkers, and collagen-deposition/fibrotic areas as well as epithelial-damaged score displayed an identical pattern compared to that of urine albumin among the three groups (all P<0.0001). In conclusion, ECSW treatment effectively protected urinary bladder from radiation-induced CC.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5883143PMC
March 2018

Melatonin attenuated brain death tissue extract-induced cardiac damage by suppressing DAMP signaling.

Oncotarget 2018 Jan 12;9(3):3531-3548. Epub 2017 Dec 12.

Division of Cardiology, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University College of Medicine, Kaohsiung, Taiwan.

We tested the hypothesis that melatonin prevents brain death (BD) tissue extract (BDEX)-induced cardiac damage by suppressing inflammatory damage-associated molecular pattern (DAMP) signaling in rats. Six hours after BD induction, levels of a DAMP component (HMGB1) and inflammatory markers (TLR-2, TLR-4, MYD88, IκB, NF-κB, IL-1β, IFN-γ, TNF-α and IL-6) were higher in brain tissue from BD animals than controls. Levels of HMGB1 and inflammatory markers were higher in BDEX-treated H9C2 cardiac myoblasts than in cells treated with healthy brain tissue extract. These increases were attenuated by melatonin but re-induced with luzindole (all < 0.001). Additional male rats ( 30) were divided into groups 1 (negative control), 2 (healthy brain tissue extract implanted in the left ventricular myocardium [LVM]), 3 (BDEX-LVM), 4 (BDEX-LVM + melatonin), and 5 (BDEX-LVM + melatonin + luzindole). Collagen deposition/fibrosis and LVM levels of MTR2, HMGB1, inflammatory markers, oxidative stress, apoptosis, mitochondrial damage and DNA damage were highest in group 3, lowest in groups 1 and 2, and higher in group 5 than in group 4. Heart function and LVM levels of MTR1 and anti-inflammatory, mitochondrial-integrity and anti-oxidative markers exhibited a pattern opposite that of the inflammatory markers in the five groups (all < 0.0001). These results indicate melatonin inhibits BDEX-induced cardiac damage by suppressing the DAMP inflammatory axis.
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http://dx.doi.org/10.18632/oncotarget.23180DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5790481PMC
January 2018

Therapeutic effects of adipose-derived mesenchymal stem cells against brain death-induced remote organ damage and post-heart transplant acute rejection.

Oncotarget 2017 Dec 30;8(65):108692-108711. Epub 2017 Sep 30.

Division of Thoracic and Cardiovascular Surgery, Department of Surgery, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University College of Medicine, Kaohsiung, Taiwan.

We tested the hypothesis that allogenic adipose-derived mesenchymal stem cells (ADMSCs) alleviated brain death (BD)-induced remote organ damage and events of post heart-transplant acute rejection. To determine the impact of BD on remote organ damage, adult-male F344 rats (n=24) were categorized sham-control (SC), BD and BD (allogenic ADMSC/1.2 × 10 cells/derived from F344 by intravenous transfusion 3 h after BD procedure). To determine the protective effect of allogenic ADMSCs, animals (n=8/each group in F344/Lewis) were categorized into groups BD-T(F344 heart transplanted into Lewis by 6h after BD), BD-T (BD induction for 6h then heart transplantation, and allogenic ADMSCs transfusion at days 1 and 5 after heart transplantation), BD-T (BD + ADMSC/1.2 × 10 cells at 3h and heart transplantation at 6h after BD) and BD-T [BD + ADMSC/1.2 × 10 cells at 3h and heart transplantation at 6h after BD, then ADMSC therapy by days 1/3]. At day 5 post procedure, liver, kidney and heart specimens showed higher molecular-cellular levels of inflammation, immune reaction, apoptosis and fibrosis in BD than in SC that were reversed in BD (all P < 0.0001). These molecular-cellular expressions and circulating/splenic levels of innate/adoptive immune cells were higher in BD-T, lowest in BD-T and higher BD-T in than BD-T, whereas heart function showed an opposite pattern among the four groups (all P < 0.001). In conclusion, ADMSCs suppressed BD-caused remote organ damage and heart-transplant rejection.
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http://dx.doi.org/10.18632/oncotarget.21433DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5752474PMC
December 2017
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