Pyrophosphate therapy prevents trauma-induced calcification in the mouse model of neurogenic heterotopic ossification.

Trauma-induced calcification is the pathological consequence of complex injuries which often affect the central nervous system and other parts of the body simultaneously. We demonstrated by an animal model recapitulating the calcification of the above condition that adrenaline transmits the stress signal of brain injury to the calcifying tissues. We have also found that although the level of plasma pyrophosphate, the endogenous inhibitor of calcification, was normal in calcifying animals, it could not counteract the acute calcification. However, externally added pyrophosphate inhibited calcification even when it was administered after the complex injuries. Our finding suggests a potentially powerful clinical intervention of calcification triggered by polytrauma injuries which has no effective treatment.

Cylindrospermopsin induces biochemical changes leading to programmed cell death in plants.

In the present study we provide cytological and biochemical evidence that the cyanotoxin cylindrospermopsin (CYN) induces programmed cell death (PCD) symptoms in two model vascular plants: the dicot white mustard (Sinapis alba) and the monocot common reed (Phragmites australis). Cytological data include chromatin fragmentation and the increase of the ratio of TUNEL-positive cells in roots, the latter being detected in both model systems studied. The strongest biochemical evidence is the elevation of the activity of several single-stranded DNA preferring nucleases-among them enzymes active at both acidic and alkaline conditions and are probably directly related to DNA breaks occurring at the initial stages of plant PCD: 80 kDa nucleases and a 26 kDa nuclease, both having dual (single- and double-stranded nucleic acid) specificity. Moreover, the total protease activity and in particular, a 53-56 kDa alkaline protease activity increases. This protease could be inhibited by PMSF, thus regarded as serine protease. Serine proteases are detected in all organs of Brassicaceae (Arabidopsis) having importance in differentiation of specialized plant tissue through PCD, in protein degradation/processing during early germination and defense mechanisms induced by a variety of biotic and abiotic stresses. However, knowledge of the physiological roles of these proteases and nucleases in PCD still needs further research. It is concluded that CYN treatment induces chromatin fragmentation and PCD in plant cells by activating specific nucleases and proteases. CYN is proposed to be a suitable molecule to study the mechanism of plant apoptosis.

3D culturing of human pluripotent stem cells-derived endothelial cells for vascular regeneration.

Rationale: Human induced pluripotent stem cell-derived endothelial cells can be candidates for engineering therapeutic vascular grafts. Methods: Here, we studied the role of three-dimensional culture on their characteristics and function both in vitro and in vivo. Results: We found that differentiated hPSC-EC can re-populate decellularized biomatrices; they remain viable, undergo maturation and arterial/venous specification. Human PSC-EC develop antifibrotic, vasoactive and anti-inflammatory properties during recellularization. In vivo, a robust increase in perfusion was detected at the engraftment sites after subcutaneous implantation of an hPSC-EC-laden hydrogel in rats. Histology confirmed survival and formation of capillary-like structures, suggesting the incorporation of hPSC-EC into host microvasculature. In a canine model, hiPSC-EC-seeded onto decellularised vascular segments were functional as aortic grafts. Similarly, we showed the retention and maturation of hiPSC-EC and dynamic remodelling of the vessel wall with good maintenance of vascular patency. Conclusions: A combination of hPSC-EC and biomatrices may be a promising approach to repair ischemic tissues.

Nucleoside-modified VEGFC mRNA induces organ-specific lymphatic growth and reverses experimental lymphedema.

Lack or dysfunction of the lymphatics leads to secondary lymphedema formation that seriously reduces the function of the affected organs and results in degradation of quality of life. Currently, there is no definitive treatment option for lymphedema. Here, we utilized nucleoside-modified mRNA encapsulated in lipid nanoparticles (LNPs) encoding murine Vascular Endothelial Growth Factor C (VEGFC) to stimulate lymphatic growth and function and reduce experimental lymphedema in mouse models. We demonstrated that administration of a single low-dose of VEGFC mRNA-LNPs induced durable, organ-specific lymphatic growth and formation of a functional lymphatic network. Importantly, VEGFC mRNA-LNP treatment reversed experimental lymphedema by restoring lymphatic function without inducing any obvious adverse events. Collectively, we present a novel application of the nucleoside-modified mRNA-LNP platform, describe a model for identifying the organ-specific physiological and pathophysiological roles of the lymphatics, and propose an efficient and safe treatment option that may serve as a novel therapeutic tool to reduce lymphedema.

Reduced Prenatal Pulmonary Lymphatic Function Is Observed in Clp1 K/K Embryos With Impaired Motor Functions Including Fetal Breathing Movements in Preparation of the Developing Lung for Inflation at Birth.

Embryonic lungs must be inflated immediately after birth to establish respiration. In addition to pulmonary surfactant, recently, we have revealed lymphatic function as a previously unknown regulator of prenatal lung compliance that prepares the embryonic lung for inflation at birth. It is well-documented that the late gestation embryo performs episodic breathing-like movements called as fetal breathing movements (FBMs), but the physiological importance of these events is not clear. Here we aimed to study the physiological role of FBMs in preparation for air inflation at birth. Clp1 K/K late gestation embryos develop a progressive loss of spinal motor neurons associated with axonal degeneration and denervation of neuromuscular junctions serving as an ideal genetic model to test the possible role of FBMs. We demonstrated that Clp1 K/K newborns show impaired motor function resulting in fatal respiratory failure after birth. Next, we showed that the alveolar septa are thicker, and the alveolar area is reduced in Clp1 K/K late gestation embryos, while the expression of molecular markers of lung development are not affected. Importantly, pulmonary lymphatic vessels are dilated and the prenatal pulmonary lymphatic function is reduced in Clp1 K/K late gestation embryos. Our results have revealed that Clp1 K/K mice show impaired motor functions including FBMs, and late gestation Clp1 K/K embryos display reduced prenatal lymphatic function and impaired lung expansion represented as thickened alveolar septa and reduced alveolar area in preparation of the developing lung for inflation at birth. These findings suggest a possible mechanism that FBMs, similarly to breathing movements after birth, stimulate prenatal lymphatic function in pulmonary collecting lymphatics lacking smooth muscle coverage to prepare the developing lung for inflation and gas exchange at birth. Moreover, these results raise the possibility that stimulating FBMs during late gestation might be an effective way to reduce the risk of the development of neonatal respiratory failure.