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Recent studies indicate that mesenchymal stromal cell (MSC) transplantation improves healing of injured and diseased skeletal muscle, although the mechanisms of benefit are poorly understood. In the present study, we investigated whether MSCs and/or their trophic factors were able to regulate matrix metalloproteinase (MMP) expression and activity in different cells of the muscle tissue. MSCs in co-culture with C2C12 cells or their conditioned medium (MSC-CM) up-regulated MMP-2 and MMP-9 expression and function in the myoblastic cells; these effects were concomitant with the down-regulation of the tissue inhibitor of metalloproteinases (TIMP)-1 and -2 and with increased cell motility. In the single muscle fiber experiments, MSC-CM administration increased MMP-2/9 expression in Pax-7(+) satellite cells and stimulated their mobilization, differentiation and fusion. The anti-fibrotic properties of MSC-CM involved also the regulation of MMPs by skeletal fibroblasts and the inhibition of their differentiation into myofibroblasts. The treatment with SB-3CT, a potent MMP inhibitor, prevented in these cells, the decrease of α-smooth actin and type-I collagen expression induced by MSC-CM, suggesting that MSC-CM could attenuate the fibrogenic response through mechanisms mediated by MMPs. Our results indicate that growth factors and cytokines released by these cells may modulate the fibrotic response and improve the endogenous mechanisms of muscle repair/regeneration., (Copyright © 2014. Published by Elsevier Inc.)

The possibility to induce myocardial regeneration by the activation of resident cardiac stem cells (CSCs) has raised great interest. However, to propose endogenous CSCs as therapeutic options, a better understanding of the complex mechanisms controlling heart morphogenesis is needed, including the cellular and molecular interactions that cardiomyocyte precursors establish with cells of the stromal compartment. In the present study, we co-cultured immature cardiomyocytes from neonatal mouse hearts with mouse bone marrow-derived mesenchymal stromal cells (MSCs) to investigate whether these cells could influence cardiomyocyte growth in vitro. We found that cardiomyocyte proliferation was enhanced by direct co-culture with MSCs compared with the single cultures. We also showed that the proliferative response of the neonatal cardiomyocytes involved the activation of Notch-1 receptor by its ligand Jagged-1 expressed by the adjacent MSCs. In fact, the cardiomyocytes in contact with MSCs revealed a stronger immunoreactivity for the activated Notch-intracellular domain (Notch-ICD) as compared with those cultured alone and this response was significantly attenuated when MSCs were silenced for Jagged-1. The presence of various cardiotropic cytokines and growth factors in the conditioned medium of MSCs underscored the contribution of paracrine mechanisms to Notch-1 up-regulation by the cardiomyocytes. In conclusions these findings unveil a previously unrecognized function of MSCs in regulating cardiomyocyte proliferation through Notch-1/Jagged-1 pathway and suggest that stromal-myocardial cell juxtacrine and paracrine interactions may contribute to the development of new and more efficient cell-based myocardial repair strategies., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

The possibility that resident myocardial progenitor cells may be re-activated by transplantation of exogenous stem cells into the post-infarcted heart has been suggested as a possible mechanism to explain the heart's functional improvement after stem cell therapy. Here we studied whether differentiation of mouse neonatal immature cardiomyocytes in vitro was influenced by mouse skeletal myoblasts C2C12, wild type or engineered to secrete the cardiotropic hormone relaxin. The cultured cardiomyocytes formed spontaneously beating clusters and temporally exhibited cardiac immunophenotypical (cKit, atrial natriuretic peptide, troponin T, connexin-43, HCN4) and electrical features (inward voltage-dependent Na(+), T- and L-type Ca(2+) currents, outward and inward K(+) currents, I(f) pacemaker current). These clusters were functionally connected through nanotubular structures and undifferentiated cardiac cells in the form of flattened stripes, bridging the clusters through connexin-43-containing gap junctions. These findings suggested the existence of long distance cell-to-cell communications among the cardiomyocyte aggregates involved in the intercellular transfer of Ca(2+) signals and organelles, likely required for coordination of myocardial differentiation. Co-presence of the myoblasts greatly increased cardiomyocyte differentiation and the amount of intercellular connections. In fact, these cells formed a structural support guiding elongation of nanotubules and stripe-like cells. The secretion of relaxin by the engineered myoblasts accelerated and enhanced the cardiomyogenic potential of the co-culture. These findings underscore the possibility that grafted myoblasts and cardiotropic factors, such as relaxin, may influence regeneration of resident immature cardiac cells, thus adding a tile to the mosaic of mechanisms involved in the functional benefits of cell transplantation for cardiac repair.

Unlabelled: The role of angiotensin II in pressure overload is still debated because notwithstanding its effects on myocyte contractility angiotensin II is not an obligatory factor for the development of hypertrophy. To define the role of angiotensin II in acute pressure overload we studied the effects of AT1 blockade (valsartan 80mg per day) on myocardial contractility, cardiac growth factor gene expression, and myocardial hypertrophy in aortic banded (60mmHg) pigs. Acute pressure overload caused an abrupt reduction of myocardial contractility, measured by the end-systolic stiffness constant, and a sharp increase in end-systolic stress which rapidly normalized (within 12h) in the placebo group. In AT1-blocked animals end-systolic stiffness constant remained significantly depressed up to 24h and end-systolic stress was still elevated up to 48h (both P<0.05 vs placebo). In both groups confocal microscopy revealed that granular staining of angiotensin II in cardiomyocyte cytoplasm disappeared after 30min of pressure overload. AT1 blockade abolished following cardiac overexpression of angiotensinogen and endothelin-1 genes as shown in RT-PCR studies and the consequent angiotensin II and endothelin-1 release in the coronary circulation. Conversely, insulin-like growth factor-I and ACE mRNA overexpression, as well as the onset of left ventricular mass increase, were not significantly affected by AT1 blockade., In Conclusion: (1) mechanical stress releases preformed angiotensin II from myocyte in vivo; (2) the AT1 blockade abolishes cardiac angiotensin II and endothelin-1 production with delayed recovery of myocardial contractility; whereas (3) the overexpression of insulin-like growth factor-I gene and the development of myocardial hypertrophy are not angiotensin II-mediated effects.

Background: Patients affected by hypoparathyroidism of variable etiology are currently treated with exogenously administered vitamin D and calcium. Human parathyroid transplantation has long been investigated as a possible mean of treating these patients to prevent long-term hypocalcemia. However, the main obstacle for this treatment is represented by tissue rejection. A reliable method to efficiently protect the transplanted tissue from rejection and to allow long-term survival of the graft is the encapsulation of tissues or cells in alginate-polylysine-alginate membranes, which were successfully used for encapsulation of islets of Langerhans. The microencapsulation of parathyroid tissue fragments or of parathyroid cells becomes, therefore, a potential approach for the successful treatment of permanent symptomatic hypoparathyroidism without pharmacological immunosuppression., Materials and Methods: We describe microencapsulation of differentiated human parathyroid cells derived from adenoma or hyperplastic glands. Long-term viability, cell growth, and parathyroid hormone production of microencapsulated cells were evaluated together with responsiveness to extracellular Ca(2+)., Results: Microencapsulated parathyroid cells maintained proliferative and differentiative properties for a long term in culture with a good response to extracellular Ca(2+) concentration., Conclusions: These findings represent a crucial step toward the construction of functional bioartificial parathyroid organoids for the treatment of hypoparathyroidism in humans., (Copyright 2001 Academic Press.)

The purpose of this study was to evaluate the early changes in sarcoplasmic reticulum (SR) function and the parallel morphological and hemodynamic modifications occurring in the heart following pressure overload. As regards SR function, we also explored the levels of acylphosphatase, an enzyme which might have a regulatory effect on the SR Ca(2+) pump by hydrolyzing the phosphorylated intermediate of this transport system. Pigs subjected to pressure overload by aortic stenosis for 6, 12, 24, 48, 72, and 96 h were compared to sham-operated controls. At each of the considered times both SR Ca(2+)-ATPase activity and Ca(2+) uptake, as well as acylphosphatase activity, were significantly enhanced in the pressure overloaded compared to the control hearts, with a maximal increase at 6 h; moreover, a positive and significant correlation was found between these parameters. The modifications in the activities of Ca(2+)-ATPase and acylphosphatase reflected an increased expression of these proteins, while phospholamban did not show significant changes in its concentration nor in its phosphorylation status. As for hemodynamic parameters, rapid changes in the left ventricular function were observed and especially the early hours following the aortic stenosis appeared to be crucial for the adjustment of heart function. The most relevant morphological finding was a focal disarrangement of the myofibrillar pattern which was very evident at 6 h, and progressively attenuated at later times. Taken together our data suggest that an early adaptation to the increased hemodynamic working overload is a consistent activation of the contractile apparatus which reflects, at least in part, an enhanced SR function. Besides the changes in Ca(2+) pump protein expression, increased acylphosphatase levels might also contribute to this effect., (Copyright 2000 Academic Press.)

21-Aminosteroids (Lazaroids), acting as free radical scavengers and as membrane stabilizers, proved to have beneficial effects in various pathological conditions. In the present study we explored the effectiveness of one of these compounds, U 74389 G, in protecting pigs myocardium against the ischemia-reperfusion damage induced by transient coronary occlusion achieved by clampling the left anterior descending coronary artery. Animals were divided into three groups: control, untreated and treated. Control animals were operated but not subjected to ischemia-reperfusion; the untreated group underwent to ischemia-reperfusion without pharmacological treatment; while the treated group received the aminosteroid (4 mg/kg) before coronary occlusion and at the time of reperfusion. Specimens of myocardial tissue and blood samples were taken for morphological and biochemical studies. In the ischemic-reperfused myocardium of the untreated animals, the dominant morphological features were neutrophil infiltration, intercellular edema and severe swelling of mitochondria. All these alterations, notably neutrophil infiltration, were attenuated by aminosteroid treatment. As for the biochemical findings, the changes in adenine nucleotides and nucleosides levels, thus the reduction of energy charge, were reversed in the treated, but not in the untreated group. Myocardial concentration of malondialdehyde, which was undetectable in the control group, was raised in all the animals after reperfusion, but this effect was significantly less marked with aminosteroid treatment. In addition, the higher myocardial content of ascorbic acid and the reduced serum potential peroxidation exhibited by the treated animals compared with untreated group indicate an enhanced antioxidant protection induced by aminosteroid administration. On the other hand, the serum levels of myoglobin, cardiac troponin I and creatine kinase MB isoenzyme suggest the ability of the aminosteroid to attenuate the modifications of membrane permeability induced by ischemia-reperfusion injury. All these results lead to the conclusion that aminosteroid treatment, at least in the conditions of the present study, is effective in reducing the morphological and biochemical alterations occurring in ischemic-reperfused myocardium., (Copyright 1997 Academic Press Limited.)