Quantitative Vibrational Dynamics of the Metal Site in a Tin Porphyrin: An IR, NRVS, and DFT Study

We used a newer, synchrotron-based, spectroscopic technique (nuclear resonance vibrational spectroscopy, NRVS) in combination with a more traditional one (infrared absorption, IR) to obtain a complete, quantitative picture of the metal center vibrational dynamics in a six-coordinated tin porphyrin. From the NRVS 119Sn site-selectivity and the sensitivity of the IR signal to 112Sn/119Sn isotope substitution, we identified the frequency of the antisymmetric stretching of the axial bonds (290 cm–1) and all the other vibrations involving Sn. Experimentally authenticated density functional theory (DFT) calculations aid the data interpretation by providing detailed normal mode descriptions for each observed vibration. These results may represent a starting point toward the characterization of the local vibrational dynamics of the metallic site in tin porphyrins and compounds with related structures. The quantitative complementariness between IR, NRVS, and DFT is emphasized

Adipose-derived mesenchymal stem cells markedly attenuate brain infarct size and improve neurological function in rats

<p>Abstract</p> <p>Background</p> <p>The therapeutic effect of adipose-derived mesenchymal stem cells (ADMSCs) on brain infarction area (BIA) and neurological status in a rat model of acute ischemic stroke (IS) was investigated.</p> <p>Methods</p> <p>Adult male Sprague-Dawley (SD) rats (n = 30) were divided into IS plus intra-venous 1 mL saline (at 0, 12 and 24 h after IS induction) (control group) and IS plus intra-venous ADMSCs (2.0 × 10⁶) (treated interval as controls) (treatment group) after occlusion of distal left internal carotid artery. The rats were sacrificed and brain tissues were harvested on day 21 after the procedure.</p> <p>Results</p> <p>The results showed that BIA was larger in control group than in treatment group (p < 0.001). The sensorimotor functional test (Corner test) identified a higher frequency of turning movement to left in control group than in treatment group (p < 0.05). mRNA expressions of Bax, caspase 3, interleukin (IL)-18, toll-like receptor-4 and plasminogen activator inhibitor-1 were higher, whereas Bcl-2 and IL-8/Gro were lower in control group than in treatment group (all p < 0.05). Western blot demonstrated a lower CXCR4 and stromal-cell derived factor-1 (SDF-1) in control group than in treatment group (all p < 0.01). Immunohistofluorescent staining showed lower expressions of CXCR4, SDF-1, von Willebran factor and doublecortin, whereas the number of apoptotic nuclei on TUNEL assay was higher in control group than in treatment group (all p < 0.001). Immunohistochemical staining showed that cellular proliferation and number of small vessels were lower but glial fibrillary acid protein was higher in control group than in treatment group (all p < 0.01).</p> <p>Conclusions</p> <p>ADMSC therapy significantly limited BIA and improved sensorimotor dysfunction after acute IS.</p

Directed Evolution of a Lysosomal Enzyme with Enhanced Activity at Neutral pH by Mammalian Cell-Surface Display

SummaryHuman β-glucuronidase, due to low intrinsic immunogenicity in humans, is an attractive enzyme for tumor-specific prodrug activation, but its utility is hindered by low activity at physiological pH. Here we describe the development of a high-throughput screening procedure for enzymatic activity based on the stable retention of fluorescent reaction product in mammalian cells expressing properly folded glycoproteins on their surface. We utilized this procedure on error-prone PCR and saturation mutagenesis libraries to isolate β-glucuronidase tetramers that were up to 60-fold more active (kcat/Km) at pH 7.0 and were up to an order of magnitude more effective at catalyzing the conversion of two structurally disparate glucuronide prodrugs to anticancer agents. The screening procedure described here can facilitate investigation of eukaryotic enzymes requiring posttranslational modifications for biological activity

Myocardium-derived conditioned medium improves left ventricular function in rodent acute myocardial infarction

<p>Abstract</p> <p>Background</p> <p>We investigated whether myocardium-derived conditioned medium (MDCM) is effective in preserving left ventricular (LV) function in a rat acute myocardial infarction (AMI) model.</p> <p>Methods</p> <p>Adult male Sprague-Dawley (SD) rats (n = 36) randomized to receive either left coronary artery ligation (AMI induction) or thoracotomy only (sham procedure) were grouped as follows (n = 6 per group): Group I, II, and III were sham-controls treated by fresh medium, normal rat MDCM, and infarct-related MDCM, respectively. Group IV, V, and VI were AMI rats treated by fresh medium, normal MDCM, and infarct-related MDCM, respectively. Either 75 μL MDCM or fresh medium was administered into infarct myocardium, followed by intravenous injection (3 mL) at postoperative 1, 12, and 24 h.</p> <p>Results</p> <p>In vitro studies showed higher phosphorylated MMP-2 and MMP-9, but lower α-smooth muscle actin and collagen expressions in neonatal cardiac fibroblasts treated with MDCM compared with those in the cardiac fibroblasts treated with fresh medium (all p < 0.05). Sirius-red staining showed larger collagen deposition area in LV myocardium in Group IV than in other groups (all p < 0.05). Stromal cell-derived factor-1α and CXCR4 protein expressions were higher in Group VI than in other groups (all p < 0.05). The number of von Willebrand factor- and BrdU-positive cells and small vessels in LV myocardium as well as 90-day LV ejection fraction were higher, whereas oxidative stress was lower in Group VI than in Group IV and Group V (all p < 0.05).</p> <p>Conclusion</p> <p>MDCM therapy reduced cardiac fibrosis and oxidative stress, enhanced angiogenesis, and preserved 90-day LV function in a rat AMI model.</p

Levels and values of circulating endothelial progenitor cells, soluble angiogenic factors, and mononuclear cell apoptosis in liver cirrhosis patients

BACKGROUND: The roles of circulating endothelial progenitor cell (EPC) and mononuclear cell apoptosis (MCA) in liver cirrhosis (LC) patients are unknown. Moreover, vascular endothelial growth factor (VEGF) and stromal cell-derived factor (SDF)-1α are powerful endogenous substances enhancing EPC migration into circulation. We assessed the level and function of EPCs [CD31/CD34 (E(1)), KDR/CD34 (E(2)), CXCR4/CD34 (E(3))], levels of MCA, VEGF and SDF-1α in circulation of LC patients. METHODS: Blood sample was prospectively collected once for assessing EPC level and function, MCA, and plasma levels of VEGF and SDF-1α using flow cytometry and enzyme-linked immunosorbent assay (ELISA), respectively, in 78 LC patients and 25 age- and gender-matched healthy controls. RESULTS: Number of EPCs (E(1), E(2), E(3)) was lower (all p < 0.0001), whereas SDF-1α level and MCA were higher (p < 0.001) in study patients compared with healthy controls. Number of EPCs (E(2), E(3)) was higher but MCA was lower (all p < 0.05) in Child's class A compared with Child's class B and C patients, although no difference in VEGF and SDF-1α levels were noted among these patients. Chronic hepatitis B and esophageal varices bleeding were independently, whereas chronic hepatitis C, elevated aspartate aminotransferase (AST), and decompensated LC were inversely and independently correlated with circulating EPC level (all p < 0.03). Additionally, angiogenesis and transwell migratory ability of EPCs were reduced in LC patients than in controls (all p < 0.001). CONCLUSION: The results of this study demonstrated that level, angiogenic capacity, and function of circulating EPCs were significantly reduced, whereas plasma levels of SDF-1α and circulating MCA were substantially enhanced in cirrhotic patients

TEX11 is mutated in infertile men with azoospermia and regulates genome-wide recombination rates in mouse

Genome‐wide recombination is essential for genome stability, evolution, and speciation. Mouse Tex11, an X‐linked meiosis‐specific gene, promotes meiotic recombination and chromosomal synapsis. Here, we report that TEX11 is mutated in infertile men with non‐obstructive azoospermia and that an analogous mutation in the mouse impairs meiosis. Genetic screening of a large cohort of idiopathic infertile men reveals that TEX11 mutations, including frameshift and splicing acceptor site mutations, cause infertility in 1% of azoospermic men. Functional evaluation of three analogous human TEX11 missense mutations in transgenic mouse models identified one mutation (V748A) as a potential infertility allele and found two mutations non‐causative. In the mouse model, an intronless autosomal Tex11 transgene functionally substitutes for the X‐linked Tex11 gene, providing genetic evidence for the X‐to‐autosomal retrotransposition evolution phenomenon. Furthermore, we find that TEX11 protein levels modulate genome‐wide recombination rates in both sexes. These studies indicate that TEX11 alleles affecting expression level or substituting single amino acids may contribute to variations in recombination rates between sexes and among individuals in humans.Howard Hughes Medical Institute (Award)National Institutes of Health (U.S.) (NIH/NIGMS grant R01GM076327

Intra-coronary administration of tacrolimus markedly attenuates infarct size and preserves heart function in porcine myocardial infarction

BACKGROUND: We test the hypothesis that intra-coronary tacrolimus administration can limit infarct size and preserve left ventricular ejection fraction (LVEF) after acute myocardial infarction (AMI) through ligating left anterior descending coronary artery (LAD) in mini-pigs. METHODS: Twelve male mini-pigs were randomized into AMI-saline (MI-only) group and AMI-tacrolimus (MI-Tac) group that received intra-coronary saline (3.0 mL) and tacrolimus (0.5 mg in 2.5 mL saline) injection, respectively, beyond site of ligation 30 minutes after LAD occlusion. RESULTS: Larger infarct area was noted in MI-only group (p < 0.001). Inflammatory biomarkers at protein [oxidative stress, tumor necrotic factor-α, nuclear factor-κB], gene (matrix metalloproteinase-9, plasminogen activator inhibitor-1), and cellular (CD40+, CD68+ inflammatory cells) levels were remarkably higher in MI-only animals (p < 0.01). Conversely, anti-inflammatory biomarkers at gene level (Interleukin-10), gene and protein level (endothelial nitric oxide synthase), and anti-oxidant biomarkers at both gene and protein levels [heme oxygenase 1, NAD(P)H:quinone oxidoreductase] were lower in MI-only group (p < 0.01). Number of apoptotic nuclei and apoptotic biomarkers expressions at gene and protein levels (Bax, caspase 3) were notably higher, whereas anti-apoptotic biomarkers at gene and protein levels (Bcl-2), LVEF, and fractional shortening were markedly lower in MI-only group (p < 0.001). CONCLUSION: Intra-coronary administration of tacrolimus significantly attenuated infarct size and preserved LV function

Early combined treatment with sildenafil and adipose-derived mesenchymal stem cells preserves heart function in rat dilated cardiomyopathy

<p>Abstract</p> <p>Background</p> <p>We investigated whether early combined autologous adipose-derived mesenchymal stem cell (ADMSC) and sildenafil therapy offers an additive benefit in preserving heart function in rat dilated cardiomyopathy (DCM).</p> <p>Methods</p> <p>Adult Lewis rats (n = 8 per group) were divided into group 1 (normal control), group 2 (saline-treated DCM rats), group 3 [2.0 × 10⁶ADMSC implanted into left ventricular (LV) myocardium of DCM rats], group 4 (DCM rats with sildenafil 30 mg/kg/day, orally), and group 5 (DCM rats with combined ADMSC-sildenafil). Treatment was started 1 week after DCM induction and the rats were sacrificed on day 90.</p> <p>Results</p> <p>The results showed that mitochondrial protein expressions of connexin43 and cytochrome-C were lowest in group 2, and lower in groups 3 and 4 than in group 5 (p < 0.002). Conversely, oxidative index was highest in group 2, and also higher in groups 3 and 4 than in group 5 (p < 0.0003). The mRNA expressions of interleukin (IL)-10, Gro/IL-8, endothelial nitric oxide synthase, and Bcl-2 were lowest in group 2, and lower in groups 3 and 4 compared with group 5 (p < 0.0001). The mRNA expressions of matrix metalloproteinase-9, Bax, caspase 3, and stromal-cell derived factor-1α were highest in group 2, and higher in groups 3 and 4 than in group 5 (p < 0.0004). Apoptosis and fibrosis in LV myocardium were most prominent in group 2 and higher in groups 3 and 4 than in group 5, whereas angiogenesis and LV ejection fraction were lowest in group 2 and lower in groups 3 and 4 than in group 5 (p < 0.003).</p> <p>Conclusion</p> <p>Early combined ADMSC/sildenafil is superior to either treatment alone in preserving LV function.</p

Adipose-Derived Mesenchymal Stem Cell Protects Kidneys against Ischemia-Reperfusion Injury through Suppressing Oxidative Stress and Inflammatory Reaction

<p>Abstract</p> <p>Background</p> <p>Reactive oxygen species are important mediators exerting toxic effects on various organs during ischemia-reperfusion (IR) injury. We hypothesized that adipose-derived mesenchymal stem cells (ADMSCs) protect the kidney against oxidative stress and inflammatory stimuli in rat during renal IR injury.</p> <p>Methods</p> <p>Adult male Sprague-Dawley (SD) rats (n = 24) were equally randomized into group 1 (sham control), group 2 (IR plus culture medium only), and group 3 (IR plus immediate intra-renal administration of 1.0 × 10⁶autologous ADMSCs, followed by intravenous ADMSCs at 6 h and 24 h after IR). The duration of ischemia was 1 h, followed by 72 hours of reperfusion before the animals were sacrificed.</p> <p>Results</p> <p>Serum creatinine and blood urea nitrogen levels and the degree of histological abnormalities were markedly lower in group 3 than in group 2 (all p < 0.03). The mRNA expressions of inflammatory, oxidative stress, and apoptotic biomarkers were lower, whereas the anti-inflammatory, anti-oxidative, and anti-apoptotic biomarkers were higher in group 3 than in group 2 (all p < 0.03). Immunofluorescent staining showed a higher number of CD31+, von Willebrand Factor+, and heme oxygenase (HO)-1+ cells in group 3 than in group 2 (all p < 0.05). Western blot showed notably higher NAD(P)H quinone oxidoreductase 1 and HO-1 activities, two indicators of anti-oxidative capacity, in group 3 than those in group 2 (all p < 0.04). Immunohistochemical staining showed higher glutathione peroxidase and glutathione reductase activities in group 3 than in group 2 (all p < 0.02)</p> <p>Conclusion</p> <p>ADMSC therapy minimized kidney damage after IR injury through suppressing oxidative stress and inflammatory response.</p