A selective fluorescent turn-on probe for imaging and sensing of hydrogen peroxide in living cells.

Fluorescent turn-on probes have been extensively used in disease diagnosis and research on pathological disease mechanisms because of their low background interference. Hydrogen peroxide (H2O2) plays a vital role in regulating various cellular functions. In the current study, a fluorescent probe, HCyB, based on hemicyanine and arylboronate structures, was designed to detect H2O2. HCyB reacted with H2O2 and exhibited a good linear relationship for H2O2 concentrations ranging from 15 to 50 µM and good selectivity over other species. The fluorescent detection limit was 76 nM. Moreover, HCyB exhibited less toxicity and mitochondrial-targeting abilities. HCyB was successfully used to monitor exogenous or endogenous H2O2 in mouse macrophage RAW 264.7, human skin fibroblast WS1, breast cancer cell MDA-MB-231, and human leukemia monocytic THP1 cells.

A Toolkit for Engineering Proteins in Living Cells: Peptide with a Tryptophan-Selective Ru-TAP Complex to Regioselectively Photolabel Specific Proteins.

Using a chemical approach to crosslink functionally versatile bioeffectors (such as peptides) to native proteins of interest (POI) directly inside a living cell is a useful toolbox for chemical biologists. However, this goal has not been reached due to unsatisfactory chemoselectivity, regioselectivity, and protein selectivity in protein labeling within living cells. Herein, we report the proof of concept of a cytocompatible and highly selective photolabeling strategy using a tryptophan-specific Ru-TAP complex as a photocrosslinker. Aside from the high selectivity, the photolabeling is blue light-driven by a photoinduced electron transfer (PeT) and allows the bioeffector to bear an additional UV-responsive unit. The two different photosensitivities are demonstrated by blue light-photocrosslinking a UV-sensitive peptide to POI. Our visible light photolabeling can generate photocaged proteins for subsequent activity manipulation by UV light. Cytoskeletal dynamics regulation is demonstrated in living cells via the unprecedented POI photomanipulation and proves that our methodology opens a new avenue to endogenous protein modification.

Cobalt protoporphyrin promotes human keratinocyte migration under hyperglycemic conditions.

BACKGROUND: Complete healing of diabetic wounds continues to be a clinically unmet need. Although robust therapies such as stem cell therapy and growth factor treatment are clinically applied, these treatments are costly for most diabetic wound patients. Therefore, a cheaper alternative is needed. Cobalt protoporphyrin (CoPP) has recently been demonstrated to promote tissue regeneration. In this study, the therapeutic benefits of CoPP in diabetic wound healing were examined. METHODS: An in vitro wound healing model that mimics re-epithelialization was established to examine the effect of CoPP on the migratory capability of human keratinocytes (HaCaT) in either normal glucose (NG) or high glucose (HG) media, as well as in the presence of either H2O2 or lipopolysaccharide (LPS). At the end of the migration assays, cells were collected and subjected to Western blotting analysis and immunostaining. RESULTS: HaCaT were found to migrate significantly more slowly in the HG media compared to the NG media. CoPP treatment was found to enhance cell migration in HG media, but was found to decrease cell migration and proliferation when HaCaT were cultured in NG media. CoPP treatment induced high levels of expression of Nrf-2/HO-1 and FoxO1 in HaCaT cultured in either glucose concentration, although the FoxO1 expression was found to be significantly higher in HaCaT that underwent the migration assay in NG media compared to those in HG media. The higher level of FoxO1 expression seen in CoPP-treated HaCaT cultured in NG media resulted in upregulation of CCL20 and downregulation of TGFß1. In contrast, HaCaT migrated in HG media were found to have high levels of expression of TGFß1, and low levels of expression of CCL20. Interestingly, in the presence of H2O2, CoPP-pretreated HaCaT cultured in either NG or HG media had similar expression level of Nrf-2/HO-1 and FoxO1 to each other. Moreover, the anti-apoptotic effect of CoPP pretreatment was noticed in HaCaT cultured in either glucose concentration. Additionally, CoPP pretreatment was shown to promote tight junction formation in HaCaT suffering from LPS-induced damage. CONCLUSIONS: CoPP enhances cell migratory capacity under hyperglycemic conditions, and protects cells from oxidative and LPS-induced cellular damage in HG media containing either H2O2 or LPS.

Platelet Factor 4 Binds to Vascular Proteoglycans and Controls Both Growth Factor Activities and Platelet Activation.

Platelet factor 4 (PF4) is produced by platelets with roles in both inflammation and wound healing. PF4 is stored in platelet α-granules bound to the glycosaminoglycan (GAG) chains of serglycin. This study revealed that platelet serglycin is decorated with chondroitin/dermatan sulfate and that PF4 binds to these GAG chains. Additionally, PF4 had a higher affinity for endothelial-derived perlecan heparan sulfate chains than serglycin GAG chains. The binding of PF4 to perlecan was found to inhibit both FGF2 signaling and platelet activation. This study revealed additional insight into the ways in which PF4 interacts with components of the vasculature to modulate cellular events.

Harnessing the early post-injury inflammatory responses for cardiac regeneration.

Cardiac inflammation is considered by many as the main driving force in prolonging the pathological condition in the heart after myocardial infarction. Immediately after cardiac ischemic injury, neutrophils are the first innate immune cells recruited to the ischemic myocardium within the first 24 h. Once they have infiltrated the injured myocardium, neutrophils would then secret proteases that promote cardiac remodeling and chemokines that enhance the recruitment of monocytes from the spleen, in which the recruitment peaks at 72 h after myocardial infarction. Monocytes would transdifferentiate into macrophages after transmigrating into the infarct area. Both neutrophils and monocytes-derived macrophages are known to release proteases and cytokines that are detrimental to the surviving cardiomyocytes. Paradoxically, these inflammatory cells also play critical roles in repairing the injured myocardium. Depletion of either neutrophils or monocytes do not improve overall cardiac function after myocardial infarction. Instead, the left ventricular function is further impaired and cardiac fibrosis persists. Moreover, the inflammatory microenvironment created by the infiltrated neutrophils and monocytes-derived macrophages is essential for the recruitment of cardiac progenitor cells. Recent studies also suggest that treatment with anti-inflammatory drugs may cause cardiac dysfunction after injury. Indeed, clinical studies have shown that traditional ant-inflammatory strategies are ineffective to improve cardiac function after infarction. Thus, the focus should be on how to harness these inflammatory events to either improve the efficacy of the delivered drugs or to favor the recruitment of cardiac progenitor cells.

Bioengineered human heparin with anticoagulant activity.

Heparin is a carbohydrate anticoagulant used clinically to prevent thrombosis, however impurities can limit its efficacy. Here we report the biosynthesis of heparin-like heparan sulfate via the recombinant expression of human serglycin in human cells. The expressed serglycin was also decorated with chondroitin/dermatan sulfate chains and the relative abundance of these glycosaminoglycan chains changed under different concentrations of glucose in the culture medium. The recombinantly expressed serglycin produced with 25mM glucose present in the culture medium was found to possess anticoagulant activity one-seventh of that of porcine unfractionated heparin, demonstrating that bioengineered human heparin-like heparan sulfate may be a safe next-generation pharmaceutical heparin.

Mast cells produce novel shorter forms of perlecan that contain functional endorepellin: a role in angiogenesis and wound healing.

Mast cells are derived from hematopoietic progenitors that are known to migrate to and reside within connective and mucosal tissues, where they differentiate and respond to various stimuli by releasing pro-inflammatory mediators, including histamine, growth factors, and proteases. This study demonstrated that primary human mast cells as well as the rat and human mast cell lines, RBL-2H3 and HMC-1, produce the heparan sulfate proteoglycan, perlecan, with a molecular mass of 640 kDa as well as smaller molecular mass species of 300 and 130 kDa. Utilizing domain-specific antibodies coupled with N-terminal sequencing, it was confirmed that both forms contained the C-terminal module of the protein core known as endorepellin, which were generated by mast cell-derived proteases. Domain-specific RT-PCR experiments demonstrated that transcripts corresponding to domains I and V, including endorepellin, were present; however, mRNA transcripts corresponding to regions of domain III were not present, suggesting that these cells were capable of producing spliced forms of the protein core. Fractions from mast cell cultures that were enriched for these fragments were shown to bind endothelial cells via the α(2)ß(1) integrin and stimulate the migration of cells in "scratch assays," both activities of which were inhibited by incubation with either anti-endorepellin or anti-perlecan antibodies. This study shows for the first time that mast cells secrete and process the extracellular proteoglycan perlecan into fragments containing the endorepellin C-terminal region that regulate angiogenesis and matrix turnover, which are both key events in wound healing.

Prognostic impact of myosteatosis in patients with colorectal cancer undergoing curative surgery: an updated systematic review and meta-analysis.

Background: Colorectal cancer (CRC) is a global health concern, and identifying prognostic factors can improve outcomes. Myosteatosis is fat infiltration into muscles and is a potential predictor of the survival of patients with CRC. Methods: This systematic review and meta-analysis aimed to assess the prognostic role of myosteatosis in CRC. PubMed, Embase, and Cochrane CENTRAL were searched up to 1 August 2023, for relevant studies, using combinations of the keywords CRC, myosteatosis, skeletal muscle fat infiltration, and low skeletal muscle radiodensity. Case-control, prospective, and retrospective cohort studies examining the association between myosteatosis and CRC outcomes after curative intent surgery were eligible for inclusion. Primary outcomes were overall survival (OS), disease-free survival (DFS), and cancer-specific survival (CSS). Results: A total of 10 studies with a total of 9,203 patients were included. The pooled hazard ratio (HR) for OS (myosteatosis vs. no myosteatosis) was 1.52 [95% confidence interval (CI), 1.38-1.67); for CSS, 1.67 (95% CI, 1.40-1.99); and for DFS, 1.89 (95% CI, 1.35-2.65). Conclusion: In patients with CRC undergoing curative intent surgery, myosteatosis is associated with worse OS, CSS, and DFS. These findings underscore the importance of evaluating myosteatosis in patients with CRC to improve outcomes.

Effects of C825T polymorphism of the GNB3 gene on availability of dopamine transporter in healthy volunteers--a SPECT study.

Striatal dopaminergic activity is significantly correlated with various cognitive activities, mood regulation, and even metabolic homeostasis, and is modulated by the dopamine transporter (DAT). The availability of DAT could be regulated by presynaptic autoreceptors, which are G-protein coupled receptors; however, whether functional variations in the common downstream signaling molecule, G-protein, could cause individual differences in presynaptic transporter availability remains unclear. To investigate this relationship, the DAT availability in seventy-eight healthy subjects was approximated using single photon emission computed tomography (SPECT) with [(99m)Tc] TRODAT-1, a radio-labeled form of tropan derivative for the selective labeling of DAT. The C825T single nucleotide polymorphism (SNP) (rs5443) of the beta subunit of the G-protein second messenger (GNß3) gene was genotyped, and analysis of variance showed a significant difference in striatal DAT when referenced to the entire occipital lobe among the three genotypes. Post hoc independent t tests were also performed, and showed that the striatal DAT availability of the CC genotype was higher than that of the other two genotypes. These results indicated that genetic variation in the common downstream signaling molecule of the dopamine autoreceptor could affect the functional status of the striatal dopamine system. These results together with the known role of the GNß3 gene might provide further evidence to support the common effect of the striatal dopamine system on mood and metabolic regulation.

Tumor targeting with DGEA peptide ligands: a new aromatic peptide amphiphile for imaging cancers.

We report a novel fluorescent bioprobe, tetraphenylethylene-Phe-Asp-Gly-Glu-Ala (TPE-FDGEA), and its self-assembly behavior, photophysical properties, and biocompatibility. The hydrogelator TPE-FDGEA exhibited aggregation-induced emission characteristics, which facilitated imaging of PC-3 human prostate cancer cells, thereby demonstrating the utility of such fluorescent probes for specific labeling of target cells in vitro.

Glucosamine-Based Supramolecular Nanotubes for Human Mesenchymal Cell Therapy.

Herein, we demonstrate an example of glucosamine-based supramolecular hydrogels that can be used for human mesenchymal cell therapy. We designed and synthesized a series of amino acid derivatives based on a strategy of capping d-glucosamine moiety at the C-terminus and fluorinated benzyl group at the N-terminus. From a systematic study on chemical structures, we discovered that the glucosamine-based supramolecular hydrogel [pentafluorobenzyl (PFB)-F-Glu] self-assembled with one-dimensional nanotubular structures at physiological pH. The self-assembly of a newly discovered PFB-F-Glu motif is attributed to the synergistic effect of π-π stacking and extensive intermolecular hydrogen bonding network in aqueous medium. Notably, PFB-F-Glu nanotubes are proven to be nontoxic to human mesenchymal stem cells (hMSCs) and have been shown to enhance hMSC proliferation while maintaining their pluripotency. Retaining of pluripotency capabilities provides potentially unlimited source of undifferentiated cells for the treatment of future cell therapies. Furthermore, hMSCs cultured on PFB-F-Glu are able to secrete paracrine factors that downregulate profibrotic gene expression in lipopolysaccharide-treated human skin fibroblasts, which demonstrates that PFB-F-Glu nanotubes have the potential to be used for wound healing applications. Overall, this article addresses the importance of chemical design to generate supramolecular biomaterials for stem cell therapy.

The formulation and characterization of 3D printed grafts as vascular access for potential use in hemodialysis.

Arteriovenous graft (AVG) failure continues to be a life-threatening problem in haemodialysis. Graft failure can occur if the implanted graft is not well-matched to the vasculature of the patient. Likewise, stenosis often develops at the vein-graft anastomosis, contributing to thrombosis and early graft failure. To address this clinical need, a novel ink formulation comprised of ACMO/TMPTA/TMETA for 3D printing a AVG was developed (ACMO-AVG), in which the printed AVG was biocompatible and did not induce cytotoxicity. The ease of customizing the ACMO-AVG according to different requirements was demonstrated. Furthermore, the AVG displayed similar mechanical properties to the commercially available arteriovenous ePTFE graft (ePTFE-AVG). Unlike ePTFE-AVG, the ACMO-AVG displayed excellent anti-fouling characteristics because no plasma protein adsorption and platelet adhesion were detected on the luminal surfaces after 2 h of incubation. Similarly, exposure to human endothelial cells and human vascular smooth muscle cells did not result in any cell detection on the surfaces of the ACMO-AVG. Thus, the present study demonstrates a newly developed 3D printing ink formulation that can be successfully 3D printed into a clinically applicable vascular access used for haemodialysis.

Subcellular Localization of Survivin Determines Its Function in Cardiomyocytes.

Rationale: Reducing cardiomyocyte death and enhancing their proliferation after myocardial infarction is perhaps the single largest challenge for cardiac tissue regeneration. Survivin (SVV) is the smallest member of the inhibitor of apoptosis (IAP) family but plays two important roles; inhibiting caspase-9 activation in the intrinsic apoptosis pathway, and regulating microtubule dynamics and chromosome segregation during cell division. Genetic depletion of cardiac SVV leads to incomplete cardiomyocyte division and abnormal heart development. However, the function of SVV in adult hearts after myocardial infarction remains unclear. Methods: A homozygous inducible cardiomyocyte-specific SVV knockout transgenic mouse model was established through crossbreeding SVVflox/flox and αMHC-MCM transgenic mice. Adult mice received consecutive intraperitoneal injection of tamoxifen to induce genetic removal of SVV in cardiomyocytes. A SVV overexpressing model was established via local delivery of SVV in wild-type mouse hearts. Results: We found that 30.82% of cardiomyocytes in the peri-infarct region of SVV knockout mice were apoptotic, significantly higher than the 22.18% in control mice. In addition, ejection fraction was 29.00±0.40% in knockout mice compared to 38.04±0.50% in control mice 21 days after myocardial infarction. On the contrary, locally overexpressing SVV in the heart improved cardiac functions. Unexpectedly, we found that altering the subcellular localization of SVV overexpression produced different outcomes. Overexpression of SVV in the cytoplasm decreased cardiomyocyte apoptosis, whereas overexpression of SVV in the nucleus enhanced cardiac regeneration. The ejection fraction of mice overexpressing SVV was 36.58±0.91%, significantly higher than 28.18±1.70% in the GFP control group. Apoptotic cardiomyocytes were only 4.63% in mouse overexpressing cytosolic SVV, compared to 9.31% in the GFP group, and activation of caspase-3 was also reduced. Moreover, mice overexpressing NLS-SVV exhibited a better ejection fraction (36.19±1.02%,) than GFP controls (26.69±0.75%). NLS-SVV enhanced H3P-positive cardiomyocytes in the border zone to 0.28%, compared to only 0.08% in GFP group, through interacting with Aurora B. Conclusions: We demonstrate the importance of SVV subcellular localization in regulating post-MI cardiac repair and regeneration. We hope that this will open new translational approaches through targeted delivery of SVV.

Syndecans as Cell Surface Receptors in Cancer Biology. A Focus on their Interaction with PDZ Domain Proteins.

Syndecans are transmembrane receptors with ectodomains that are modified by glycosaminoglycan chains. The ectodomains can interact with a wide variety of molecules, including growth factors, cytokines, proteinases, adhesion receptors, and extracellular matrix (ECM) components. The four syndecans in mammals are expressed in a development-, cell-type-, and tissue-specific manner and can function either as co-receptors with other cell surface receptors or as independent adhesion receptors that mediate cell signaling. They help regulate cell proliferation and migration, angiogenesis, cell/cell and cell/ECM adhesion, and they may participate in several key tumorigenesis processes. In some cancers, syndecan expression regulates tumor cell proliferation, adhesion, motility, and other functions, and may be a prognostic marker for tumor progression and patient survival. The short cytoplasmic tail is likely to be involved in these events through recruitment of signaling partners. In particular, the conserved carboxyl-terminal EFYA tetrapeptide sequence that is present in all syndecans binds to some PDZ domain-containing proteins that may function as scaffold proteins that recruit signaling and cytoskeletal proteins to the plasma membrane. There is growing interest in understanding these interactions at both the structural and biological levels, and recent findings show their high degree of complexity. Parameters that influence the recruitment of PDZ domain proteins by syndecans, such as binding specificity and affinity, are the focus of active investigations and are important for understanding regulatory mechanisms. Recent studies show that binding may be affected by post-translational events that influence regulatory mechanisms, such as phosphorylation within the syndecan cytoplasmic tail.

Injection of Human Cord Blood Cells With Hyaluronan Improves Postinfarction Cardiac Repair in Pigs.

UNLABELLED: Recent clinical trials using autologous bone marrow or peripheral blood cells to treat myocardial infarction (MI) show controversial results, although the treatment has a good safety profile. These discrepancies are likely caused by factors such as aging, systemic inflammation, and cell processing procedures, all of which might impair the regenerative capability of the cells used. Here, we tested whether injection of human cord blood mononuclear cells (CB-MNCs) combined with hyaluronan (HA) hydrogel improves cell therapy efficacy in a pig MI model. A total of 34 minipigs were divided into 5 groups: sham operation (Sham), surgically induced-MI plus injection with normal saline (MI+NS), HA only (MI+HA), CB-MNC only (MI+CB-MNC), or CB-MNC combined with HA (MI+CB-MNC/HA). Two months after the surgery, injection of MI+CB-MNC/HA showed the highest left ventricle ejection fraction (51.32%±0.81%) compared with MI+NS (42.87%±0.97%, p<.001), MI+HA (44.2%±0.63%, p<.001), and MI+CB-MNC (46.17%±0.39%, p<.001) groups. The hemodynamics data showed that MI+CB-MNC/HA improved the systolic function (+dp/dt) and diastolic function (-dp/dt) as opposed to the other experimental groups, of which the CB-MNC alone group only modestly improved the systolic function (+dp/dt). In addition, CB-MNC alone or combined with HA injection significantly decreased the scar area and promoted angiogenesis in the infarcted region. Together, these results indicate that combined CB-MNC and HA treatment improves heart performance and may be a promising treatment for ischemic heart diseases. SIGNIFICANCE: This study using healthy human cord blood mononuclear cells (CB-MNCs) to treat myocardial infarction provides preclinical evidence that combined injection of hyaluronan and human CB-MNCs after myocardial infarction significantly increases cell retention in the peri-infarct area, improves cardiac performance, and prevents cardiac remodeling. Moreover, using healthy cells to replace dysfunctional autologous cells may constitute a better strategy to achieve heart repair and regeneration.

Reloadable multidrug capturing delivery system for targeted ischemic disease treatment.

Human clinical trials of protein therapy for ischemic diseases have shown disappointing outcomes so far, mainly because of the poor circulatory half-life of growth factors in circulation and their low uptake and retention by the targeted injury site. The attachment of polyethylene glycol (PEG) extends the circulatory half-lives of protein drugs but reduces their extravasation and retention at the target site. To address this issue, we have developed a drug capture system using a mixture of hyaluronic acid (HA) hydrogel and anti-PEG immunoglobulin M antibodies, which, when injected at a target body site, can capture and retain a variety of systemically injected PEGylated therapeutics at that site. Furthermore, repeated systemic injections permit "reloading" of the capture depot, allowing the use of complex multistage therapies. This study demonstrates this capture system in both murine and porcine models of critical limb ischemia. The results show that the reloadable HA/anti-PEG system has the potential to be clinically applied to patients with ischemic diseases, who require sequential administration of protein drugs for optimal outcomes.

Biomimicking Platelet-Monocyte Interactions as a Novel Targeting Strategy for Heart Healing.

In patients who survive myocardial infarction, many go on to develop congestive heart failure (CHF). Despite ongoing efforts to develop new approaches for postinfarction therapy, there are still no effective therapeutic options available to CHF patients. Currently, the delivery of cardioprotective drugs relies entirely on passive uptake via the enhanced permeability and retention (EPR) effect which occurs in proximity to the infarction site. However, in ischemic disease, unlike in cancer, the EPR effect only exists for a short duration postinfarction and thus insufficient for meaningful cardioprotection. Splenic monocytes are recruited to the heart in large numbers postinfarction, and are known to interact with platelets during circulation. Therefore, the strategy is to exploit this interaction by developing platelet-like proteoliposomes (PLPs), biomimicking platelet interactions with circulating monocytes. PLPs show strong binding affinity for monocytes but not for endothelial cells in vitro, mimicking normal platelet activity. Furthermore, intravital multiphoton imaging shows that comparing to plain liposomes, PLPs do not aggregate on uninjured endothelium but do accumulate at the injury site 72 h postinfarction. Importantly, PLPs enhance the targeting of anti-inflammatory drug, cobalt protoporphyrin, to the heart in an EPR-independent manner, which result in better therapeutic outcome.

Injection of Peptide nanogels preserves postinfarct diastolic function and prolongs efficacy of cell therapy in pigs.

Accumulating evidence suggests that the benefits of cell therapy for cardiac repair are modest and transient due to progressive harmful cardiac remodeling as well as loss of transplanted cells. We previously demonstrated that injection of peptide nanofibers (NFs) reduces ventricular remodeling and facilitates cell retention at 1 month after acute myocardial infarction (MI) in pigs. However, it remains unclear whether these benefits still persist as the material is being degraded. In this study, 2 mL of placebo or NFs, with or without 1×10(8) mononuclear cells (MNCs), was injected into the pig myocardium after MI (n≥5 in each group), and cardiac function was assessed by echocardiography, including myocardial deformation analyses and catheterization at 3 months post-MI. Our results reveal that MNC-only injection slightly improved cardiac systolic function at 1 month post-MI, but this benefit was lost at later time points (ejection fraction: 42.0±2.3 in MI+normal saline [NS] and 43.5±1.1 in MI+MNCs). In contrast, NF-only injection resulted in improved cardiac diastolic function and reduced pathological remodeling at 3 months post-MI. Furthermore, combined injection of MNCs/NFs provided a greater and longer term cardiac performance (52.1±1.2 in MI+MNCs/NFs, p<0.001 versus MI+NS and MI+MNCs) and 11.3-fold transplanted cell retention. We also found that about 30% NFs remained at 3 months after injection; however, endogenous myofibroblasts were recruited to the NF-injected microenvironment to replace the degraded NFs and preserved cardiac dimensions and mechanics. In conclusion, we demonstrated that injection of NFs contributes to preservation of ventricular mechanical integrity and sustains MNC efficacy at 3 months postinjection.

The time window for therapy with peptide nanofibers combined with autologous bone marrow cells in pigs after acute myocardial infarction.

BACKGROUND: We previously showed that injection of peptide nanofibers (NF) combined with autologous bone marrow mononuclear cells (MNC) immediately after coronary artery ligation improves cardiac performance in pigs. To evaluate the clinical feasibility, this study was performed to determine the therapeutic time window for NF/MNC therapy in acute myocardial infarction (MI). METHODS AND RESULTS: A total of 45 adult minipigs were randomly grouped into 7 groups: sham or MI plus treatment with NS (normal saline), or NF or MNC alone at 1 day (1D) post-MI, or NF/MNC at 1, 4, or 7 days post-MI (N≥6). Cardiac function was assessed by echocardiography and ventricular catheterization. Compared with the NS control, pigs treated with NF/MNC at 1 day post-MI (NF/MC-1D) had the greatest improvement in left ventricle ejection fraction (LVEF; 55.1±1.6%; P<0.01 vs. NS) 2 months after MI. In contrast, pigs treated with either NF/MNC-4D or NF/MNC-7D showed 48.9±0.8% (P<0.05 vs. NS) and 43.5±2.3% (n.s. vs. NS) improvements, respectively. The +dP/dt and -dP/dt, infarct size and interstitial collagen content were also improved in the NF/MNC-1D and -4D groups but not in the -7D group. Mechanistically, MNC quality and the states of systemic inflammation and damaged heart tissue influence the therapeutic efficiency of NF/MNC therapy, as revealed by another independent study using 16 pigs. CONCLUSIONS: Injection of NF/MNC at 1 or 4 days, but not at 7 days post-MI, improves cardiac performance and prevents ventricular remodeling, confirming the importance of early intervention when using this therapy for acute MI.

Transcriptional complexity of the HSPG2 gene in the human mast cell line, HMC-1.

The mammalian HSPG2 gene encodes the proteoglycan protein core perlecan, which has important functions in biology including cell adhesion via integrins, binding to the extracellular matrix via various protein-protein interactions and binding of growth factors via the heparan sulfate chains decorating the N-terminal domain I. Here we show that, in the human mast cell line HMC-1, the transcription of this gene results in a population of mRNA that is processed in such a way to provide a relative increase of transcripts corresponding to domain V or the C-terminus compared to transcripts from either domain III or the N-terminal domain I. This paper also presents evidence of splicing of the HSPG2 gene in HMC-1 cells at exons 2/3 and after comparing this sequence with those published in various databases, a model is postulated to explain what might be happening in these cells with regard to the transcription of the HSPG2 gene. As domain V of perlecan contains the α2ß1 integrin binding site that modulates angiogenesis, we hypothesize that the transcriptional control of the HSPG2 gene in mast cells to synthesize these transcripts supports their stimulatory and specific role in wound healing and tissue regeneration.