Melatonin and mesenchymal stem cells co-administration alleviates chronic obstructive pulmonary disease via modulation of angiogenesis at the vascular-alveolar unit.

Chronic obstructive pulmonary disease (COPD) is considered a severe disease mitigating lung physiological functions with high mortality outcomes, insufficient therapy, and pathophysiology pathways which is still not fully understood. Mesenchymal stem cells (MSCs) derived from bone marrow play an important role in improving the function of organs suffering inflammation, oxidative stress, and immune reaction. It might also play a role in regenerative medicine, but that is still questionable. Additionally, Melatonin with its known antioxidative and anti-inflammatory impact is attracting attention nowadays as a useful treatment. We hypothesized that Melatonin may augment the effect of MSCs at the level of angiogenesis in COPD. In our study, the COPD model was established using cigarette smoking and lipopolysaccharide. The COPD rats were divided into four groups: COPD group, Melatonin-treated group, MSC-treated group, and combined treated group (Melatonin-MSCs). We found that COPD was accompanied by deterioration of pulmonary function tests in response to expiratory parameter affection more than inspiratory ones. This was associated with increased Hypoxia inducible factor-1α expression and vascular endothelial growth factor level. Consequently, there was increased CD31 expression indicating increased angiogenesis with massive enlargement of airspaces and thinning of alveolar septa with decreased mean radial alveolar count, in addition to, inflammatory cell infiltration and disruption of the bronchiolar epithelial wall with loss of cilia and blood vessel wall thickening. These findings were improved significantly when Melatonin and bone marrow-derived MSCs were used as a combined treatment proving the hypothesized target that Melatonin might augment MSCs aiming at vascular changes.

Synergistic cardioprotective effects of melatonin and deferoxamine through the improvement of ferritinophagy in doxorubicin-induced acute cardiotoxicity.

Ferritinophagy is one of the most recent molecular mechanisms affecting cardiac function. In addition, it is one of the pathways by which doxorubicin, one of the anticancer drugs commonly used, negatively impacts the cardiac muscle, leading to cardiac function impairment. This side effect limits the use of doxorubicin. Iron chelators play an important role in hindering ferritinophagy. Antioxidants can also impact ferritinophagy by improving oxidative stress. In this study, it was assumed that the antioxidant function of melatonin could promote the action of deferoxamine, an iron chelator, at the level of ferritinophagy. A total of 42 male Wistar rats (150-200 g) were divided into seven groups (n = 6) which consisted of group I: control normal, group II: doxorubicin (Dox), group III: melatonin (Mel), group IV: deferoxamine (Des), group V: Mel + Dox, group VI: Des + Dox, and group VII: Mel + Des + Dox. Groups III, V and VII were orally pretreated with melatonin 20 mg/kg/day for 7 days. Groups IV, VI and VII were treated with deferoxamine at a 250 mg/kg/dose once on D4 before Dox was given. Doxorubicin was given at a 20 mg/kg ip single dose. On the 8th day, the rats were lightly anaesthetized for electrocardiography analysis and echocardiography. Serum samples were collected and then sacrificed for tissue sampling. The following biochemical assessments were carried out: PCR of NCOA4, IREB2, FTH1, SLC7A11, and GPX4; and ELISA for serum cTnI, serum transferrin, tissue GSH, and malondialdehyde. In addition, histopathological assessment of heart injury; immunostaining of caspase-3, Bax, and Bcl2; and physiological function assessment by ECG and ECHO were carried out. Doxorubicin-induced acute significant cardiac injury with increased ferritinophagy and apoptosis responded to single and combined prophylactic treatment, in which the combined treatment showed mostly the best results. In conclusion, using melatonin as an antioxidant with an iron chelator, deferoxamine, could hinder the hazardous cardiotoxic effect of doxorubicin. However, further studies are needed to detect the impact of higher doses of melatonin and deferoxamine with a prolonged treatment period.

Generation of transgene-free induced pluripotent stem cells from cardiac fibroblasts of goat embryos.

Induced pluripotent stem cells (iPSCs) hold a great potential for therapeutic regenerative medicine. The aim of this study was to generate induced pluripotent stem cells from goat embryonic cardiac tissue derived fibroblasts. The isolated cardiac fibroblasts from the cardiac tissue of goat embryos were positive for alfa smooth muscle actin, vimentin and discoidin domain receptor2. From these cells, we generated transgene free iPSCs using piggyBac transposons / transposase using five transcription factors (Oct4, Sox2, Klf, Myc and Lin 28). The generated iPSCs were SSEA1, SSEA4 and Oct4 positive. They were cultured on neofeeders using 20% Serum replacement - IMDM with bFGF. They could form cystic and compact embryoid bodies that showed differentiated ectodermal and mesodermal like cells when cultured using 20% FBS-IMDM without bFGF. The iPSCs, generated in the frame of this approach were produced without the use of integrating virus and the reprogramming transgenes were removed at the end of the process. Though there were limitations in the approach used, a substantial sign of reprogramming was obtained.

The Interaction between Adult Cardiac Fibroblasts and Embryonic Stem Cell-Derived Cardiomyocytes Leads to Proarrhythmic Changes in In Vitro Cocultures.

Transplantation of stem cell-derived cardiomyocytes is one of the most promising therapeutic approaches after myocardial infarction, as loss of cardiomyocytes is virtually irreversible by endogenous repair mechanisms. In myocardial scars, transplanted cardiomyocytes will be in immediate contact with cardiac fibroblasts. While it is well documented how the electrophysiology of neonatal cardiomyocytes is modulated by cardiac fibroblasts of the same developmental stage, it is unknown how adult cardiac fibroblasts (aCFs) affect the function of embryonic stem cell-derived cardiomyocytes (ESC-CMs). To investigate the effects of aCFs on ESC-CM electrophysiology, we performed extra- and intracellular recordings of murine aCF-ESC-CM cocultures. We observed that spontaneous beating behaviour was highly irregular in aCF-ESC-CM cocultures compared to cocultures with mesenchymal stem cells (coefficient of variation of the interspike interval: 40.5 ± 15.2% versus 9.3 ± 2.0%, p = 0.008) and that action potential amplitude and maximal upstroke velocity (V max) were reduced (amplitude: 52.3 ± 1.7 mV versus 65.1 ± 1.5 mV, V max: 7.0 ± 1.0 V/s versus 36.5 ± 5.3 V/s), while action potential duration (APD) was prolonged (APD50: 25.6 ± 1.0 ms versus 16.8 ± 1.9 ms, p < 0.001; APD90: 52.2 ± 1.5 ms versus 43.3 ± 3.3 ms, p < 0.01) compared to controls. Similar changes could be induced by aCF-conditioned medium. We conclude that the presence of aCFs changes automaticity and induces potentially proarrhythmic changes of ESC-CM electrophysiology.