Pulmonary arterial hypertension in children with congenital heart disease: a deeper look into the role of endothelial progenitor cells and circulating endothelial cells to assess disease severity.

Endothelial progenitor cells and circulating endothelial cells have been proposed as useful markers of severity and disease progression in certain vascular diseases, including pulmonary arterial hypertension. Our study focused on evaluating the levels of circulating endothelial progenitor cells and circulating endothelial cells in patients with congenital left-to-right shunts and pulmonary hypertension undergoing definitive repair. Endothelial progenitor cells (identified by simultaneous co-expression of CD45dim, CD34 + and KDR2 + surface antibodies) and circulating endothelial cells (identified by simultaneous co-expression of inherent antibodies CD45-, CD31+, CD146 + and CD105+) were prospectively measured in seventy-four children (including children with Down syndrome), median age six years (2.75-10), with clinically significant left-to-right shunts undergoing transcatheter or surgical repair and compared to thirty healthy controls. Endothelial progenitor cells and, particularly, circulating endothelial cells were significantly higher in children with heart disease and pulmonary arterial hypertension when compared to controls. Endothelial progenitor cells showed significant correlation with pulmonary vascular resistance index when measured both systemically (r = 0.259; p = 0.026) and in the superior vena cava (r = 0.302; p = 0.009). Children with Down syndrome showed a stronger correlation between systemic cellularity and pulmonary vascular resistance index (r = 0.829; p = 0.002). Endothelial progenitor cells were reduced along their transit through the lung, whereas circulating endothelial cells did not suffer any modification across the pulmonary circulation. In children with yet to be repaired left-to-right shunts, endothelial progenitor cells and circulating endothelial cell counts are increased compared to healthy subjects.

Senescent CD4+CD28null cells are increased in chronic hyperuricemia, show aberrant effector phenotypes, and are reversed after allopurinol therapy: a proof-of-concept pilot study.

To characterize CD4+CD28null cells in chronic hyperuricemia and investigate whether allopurinol could restore CD28 expression and the balance of T helper phenotypes. Asymptomatic individuals with chronic hyperuricemia and ultrasonographic findings evocative of urate deposition in the joints. Age- and gender-matched normouricemic individuals were also studied. Oral allopurinol at 150 mg/day for 4 weeks, followed by 300 mg/day through week 12. Color-flow cytometry on peripheral blood mononuclear cells (PBMC) with antibodies against CD4, CD28, T-bet (Th1), GATA-3 (Th2), and RORγt (Th17). Six patients (five men, median age of 53 years) and seven controls were studied. At baseline, hyperuricemic patients had more CD4+CD28null/CD4+ cells than normouricemic subjects (36.8% vs. 6.1%; p = 0.001), with a predominance of T-bet+ cells (98.5% vs. 6.6%; p = 0.001) and few RORγt+ cells (0.7% vs. 89.4%; p = 0.014). In hyperuricemic patients, the number of CD4+ cells/10,000 PBMC was similar before and after allopurinol (3378 vs. 3954; p = 0.843). Conversely, CD4+CD28null cells decreased from 36.8% (23.0-43.7) to 15.8% (4.7-28.1; p = 0.031). CD4+CD28nullT-bet+ cells decreased from 98.5% (95.0-99.4) to 88.3% (75.2-98.9; p = 0.062), CD4+CD28nullGATA-3+ cells increased from 0% (0-4.0) to 2.8% (0.1-15.6; p = 0.156), and CD4+CD28nullRORγt+ cells increased from 0.7% (0.4-7.0) to 4.5% (1.3-28.1; p = 0.031). The CD4+CD28null cell subset is abnormally expanded in chronic hyperuricemia, despite the absence of overt urate-related disease. Allopurinol may partially restore CD28 expression on CD4+ cells while enhancing the homeostatic balance of T helper phenotypes. ClinicalTrials.gov, number NCT04012294.

Improved Efficiency of Cardiomyocyte-Like Cell Differentiation from Rat Adipose Tissue-Derived Mesenchymal Stem Cells with a Directed Differentiation Protocol.

Cell-based therapy has become a resource for the treatment of cardiovascular diseases; however, there are some conundrums to achieve. In vitro cardiomyocyte generation could be a solution for scaling options in clinical applications. Variability on cardiac differentiation in previously reported studies from adipose tissue-derived mesenchymal stem cells (ASCs) and the lack of measuring of the cardiomyocyte differentiation efficiency motivate the present study. Here, we improved the ASC-derived cardiomyocyte-like cell differentiation efficiency with a directed cardiomyocyte differentiation protocol: BMP-4 + VEGF (days 0-4) followed by a methylcellulose-based medium with cytokines (IL-6 and IL-3) (days 5-21). Cultures treated with the directed cardiomyocyte differentiation protocol showed cardiac-like cells and "rosette-like structures" from day 7. The percentage of cardiac troponin T- (cTnT-) positive cells was evaluated by flow cytometry to assess the cardiomyocyte differentiation efficiency in a quantitative manner. ASCs treated with the directed cardiomyocyte differentiation protocol obtained a differentiation efficiency of up to 44.03% (39.96%±3.78) at day 15 without any enrichment step. Also, at day 21 we observed by immunofluorescence the positive expression of early, late, and cardiac maturation differentiation markers (Gata-4, cTnT, cardiac myosin heavy chain (MyH), and the sarcoplasmic/endoplasmic reticulum Ca2+ ATPase (SERCa2)) in cultures treated with the directed cardiomyocyte differentiation protocol. Unlike other protocols, the use of critical factors of embryonic cardiomyogenesis coupled with a methylcellulose-based medium containing previously reported cardiogenic cytokines (IL-6 and IL-3) seems to be favorable for in vitro cardiomyocyte generation. This novel efficient culture protocol makes ASC-derived cardiac differentiation more efficient. Further investigation is needed to identify an ASC-derived cardiomyocyte surface marker for cardiac enrichment.

HDL-Mediated Lipid Influx to Endothelial Cells Contributes to Regulating Intercellular Adhesion Molecule (ICAM)-1 Expression and eNOS Phosphorylation.

Reverse cholesterol transport (RCT) is considered as the most important antiatherogenic role of high-density lipoproteins (HDL), but interventions based on RCT have failed to reduce the risk of coronary heart disease. In contrast to RCT, important evidence suggests that HDL deliver lipids to peripheral cells. Therefore, in this paper, we investigated whether HDL could improve endothelial function by delivering lipids to the cells. Internalization kinetics using cholesterol and apolipoprotein (apo) AI fluorescent double-labeled reconstituted HDL (rHDL), and human dermal microvascular endothelial cells-1 (HMEC-1) showed a fast cholesterol influx (10 min) and a slower HDL protein internalization as determined by confocal microscopy and flow cytometry. Sphingomyelin kinetics overlapped that of apo AI, indicating that only cholesterol became dissociated from rHDL during internalization. rHDL apo AI internalization was scavenger receptor class B type I (SR-BI)-dependent, whereas HDL cholesterol influx was independent of SR-BI and was not completely inhibited by the presence of low-density lipoproteins (LDL). HDL sphingomyelin was fundamental for intercellular adhesion molecule-1 (ICAM-1) downregulation in HMEC-1. However, vascular cell adhesion protein-1 (VCAM-1) was not inhibited by rHDL, suggesting that components such as apolipoproteins other than apo AI participate in HDL's regulation of this adhesion molecule. rHDL also induced endothelial nitric oxide synthase eNOS S1177 phosphorylation in HMEC-1 but only when the particle contained sphingomyelin. In conclusion, the internalization of HDL implies the dissociation of lipoprotein components and a SR-BI-independent fast delivery of cholesterol to endothelial cells. HDL internalization had functional implications that were mainly dependent on sphingomyelin. These results suggest a new role of HDL as lipid vectors to the cells, which could be congruent with the antiatherogenic properties of these lipoproteins.

Characterization of immortalized human dermal microvascular endothelial cells (HMEC-1) for the study of HDL functionality.

BACKGROUND: Primary cultures endothelial cells have been used as models of endothelial related diseases such atherosclerosis. Biological behavior of primary cultures is donor-dependent and data could not be easily reproducible; endothelial cell lines are emerging options, particularly, human dermal microvascular endothelial cells (HMEC-1), that should be validated to substitute primary cultures for the study of HDL functions. METHODS: Morphology, size and granularity of cells were assessed by phase contrast microscopy and flow cytometry of HMEC-1. The adhesion molecules, ICAM-1and VCAM-1 after TNF-α stimulation, and endothelial markers CD105 endoglin, as well as HDL receptor SR-BI were determined by flow cytometry. Internalization of HDL protein was demonstrated by confocal microscopy using HDL labeled with Alexa Fluor 488. HUVECs were used as reference to compared the characteristics with HMEC-1. RESULTS: HMEC-1 and HUVEC had similar morphologies, size and granularity. HMEC-1 expressed endothelial markers as HUVECs, as well as functional SR-B1 receptor since the cell line was able to internalize HDL particles. HMEC-1 effectively increased ICAM-1 and VCAM-1 expression after TNF-α stimulation. HUVECs showed more sensibility to TNF-α stimulus but the range of ICAM-1 and VCAM-1 expression was less homogeneous than in HMEC-1, probably due to biological variation of the former. Finally, the expression of adhesion molecules in HMEC-1 was attenuated by co-incubation with HDL. CONCLUSION: HMEC-1 possess characteristics of endothelial cells, similar to HUVECs, being a cell line suitable to evaluate the functionality of HDL vis-à-vis the endothelium.

Effect of fresh frozen plasma on the in vitro activation of U937 monocytes: a potential role for the age of blood donors and their underlying cytokine profile.

BACKGROUND: Fresh frozen plasma (FFP) administration may increase the risk of nosocomial infections in parallel with the development of immune modulation. This could be driven by soluble mediators, possibly influencing the in vitro activation of human U937 monocyte cells, in a manner dependent on the age of the donors. METHODS: FFP donors were stratified into groups of 19-30 years, 31-40 years or 41-50 years, and U937 cells were cultured with FFP (alone or plus lipopolysaccharide-LPS) for 24 h. Both in FFP and supernatants, TNF, IL-1ß, IL-6, and IL-10 levels were measured by ELISA. Additionally, CD11B, TLR2, and CASP3 gene expression were measured by qtPCR in U937 cells. Total phagocytic activity was also assayed. RESULTS: Elevated IL-10, but low TNF and IL-1ß levels were measured in FFP from individuals aged 19-40 years, whereas in individuals aged 41-50 years FFP were characterized by equalized TNF and IL-10 levels. Elevated IL-6 levels were found in all FFP samples, especially in those from the oldest individuals. FFP stimulation was associated with striking modifications in cytokine production in an age-dependent way. Exposure to FFP attenuates the response to LPS. TLR2 and CD11B expression were enhanced regardless of the age of plasma donors, although CASP3 expression was increased only when FFP from individuals aged 19-40 years were tested. Phagocytosis decreased after exposure to FFP regardless of donor age. CONCLUSION: Our results suggest that soluble mediators in FFP may modulate the functioning of monocytes. Interestingly, this effect appears to be partially influenced by the age of donors.

Type III Interferons in Systemic Lupus Erythematosus: Association Between Interferon λ3, Disease Activity, and Anti-Ro/SSA Antibodies.

OBJECTIVE: The aim of this study was to assess associations between serum type III (λ) interferons (IFN-λ) and disease activity in systemic lupus erythematosus (SLE). METHODS: Serum levels of IFN-λ1, IFN-λ2, and IFN-λ3 were measured in 93 SLE patients and 67 healthy individuals. The associations with overall disease activity, organ-specific damage, and SLE-related antibodies were assessed. RESULTS: Median IFN-λ1 levels were 0 pg/mL (range, 0-510 pg/mL) and 0 pg/mL (0-171 pg/mL; P = 0.814) in SLE patients and control subjects, respectively. These figures were 0 pg/mL (0-28 pg/mL) and 0 pg/mL (0-43 pg/mL; P = 0.659) for IFN-λ2, as well as 83 pg/mL (0-965 pg/mL) and 42 pg/mL (0-520 pg/mL; P = 0.002) for IFN-λ3, respectively. According to the Systemic Lupus Erythematosus Disease Activity Index categories, IFN-λ3 levels were 44 pg/mL (0-158 pg/mL) in quiescent, 117 pg/mL (0-344 pg/mL) in mild, 79 pg/mL (0-965 pg/mL) in moderate, and 78 pg/mL (0-329 pg/mL) in severe disease, with the highest levels found in patients with serosal or cutaneous involvement. In line with this, IFN-λ3 levels were inversely correlated with C3 (ρ = -0.44; 95% confidence interval, -0.62 to -0.20; P = 0.0003) and C4 (ρ = -0.40; 95% confidence interval, -0.59 to -0.15; P = 0.0001) complement proteins. In addition, higher IFN-λ3 levels were found in patients positive for anti-Ro/SSA antibodies than in those negative for that antibody (122 pg/mL [0-965 pg/mL] vs. 0 pg/mL [0-165 pg/mL]; P = 0.001). The concentration of IFN-λ3 also was higher in patients receiving glucocorticoids (104 pg/mL [0-965 pg/mL] vs. 30 pg/mL [0-165 pg/mL]; P = 0.009), and a dose-related effect was observed. CONCLUSIONS: Interferon λ3, a subtype of type III IFNs, is associated with the extent of lupus activity, in particular with active serosal and cutaneous disease. This association could be mechanistically related to anti-Ro/SSA antibodies.

Type-III interferons and rheumatoid arthritis: Correlation between interferon lambda 1 (interleukin 29) and antimutated citrullinated vimentin antibody levels.

AIM: To assess serum type III or lambda (λ) interferons (IFN) levels and its clinical and laboratory associations in rheumatoid arthritis (RA). METHODS: A cross-sectional study including 43 patients with RA (86% females; age 45.3 ± 10.3 years) and 43 healthy individuals was performed. Clinical data including disease activity, acute-phase reactants, rheumatoid factor and anticyclic citrullinated peptide (anti-CCP) antibodies were collected. Serum IFNλ1, IFNλ2, IFNλ3, CXCL8 and anti-mutated citrullinated vimentin (anti-MCV) antibody levels were measured. RESULTS: Patients with RA had higher IFNλ1 (113.5 ± 118.6 pg/mL versus 55.9 ± 122.3 pg/mL; p < 0.0001) and IFNλ2 (245.4 ± 327.7 pg/mL versus 5.1 ± 11.0 pg/mL; p = 0.009) levels than controls, but not IFNλ3 levels. Notably, IFNλ1 levels were found to be higher in both patients with active disease (124.9 ± 135.9 pg/mL; p < 0.001) and quiescent disease (99.0 ± 93.7 pg/mL; p < 0.01), while IFNλ2 levels were higher only in patients with active disease (264.0 ± 356.1 pg/mL; p = 0.02). A noteworthy association between serum IFNλ1 levels and anti-MCV antibody titers (Spearman's rho coefficient 0.36, 95% CI 0.36 to 0.61; p = 0.02) was observed. CONCLUSION: Serum IFNλ1 and IFNλ2 levels are abnormally elevated in patients with RA and the former are linearly associated with circulating anti-MCV antibody levels. These results may place type-III IFN as an attractive new therapeutic target in RA.

Effect of fresh frozen plasma on the in vitro activation of U937 monocytes: a potential role for the age of blood donors and their underlying cytokine profile

BACKGROUND: Fresh frozen plasma (FFP) administration may increase the risk of nosocomial infections in parallel with the development of immune modulation. This could be driven by soluble mediators, possibly influencing the in vitro activation of human U937 monocyte cells, in a manner dependent on the age of the donors. METHODS: FFP donors were stratified into groups of 19-30 years, 31-40 years or 41-50 years, and U937 cells were cultured with FFP (alone or plus lipopolysaccharide-LPS) for 24 h. Both in FFP and supernatants, TNF, IL-1ß, IL-6, and IL-10 levels were measured by ELISA. Additionally, CD11B, TLR2, and CASP3 gene expression were measured by qtPCR in U937 cells. Total phagocytic activity was also assayed. RESULTS: Elevated IL-10, but low TNF and IL-1ß levels were measured in FFP from individuals aged 19-40 years, whereas in individuals aged 41-50 years FFP were characterized by equalized TNF and IL-10 levels. Elevated IL-6 levels were found in all FFP samples, especially in those from the oldest individuals. FFP stimulation was associated with striking modifications in cytokine production in an age-dependent way. Exposure to FFP attenuates the response to LPS. TLR2 and CD11B expression were enhanced regardless of the age of plasma donors, although CASP3 expression was increased only when FFP from individuals aged 19-40 years were tested. Phagocytosis decreased after exposure to FFP regardless of donor age. CONCLUSION: Our results suggest that soluble mediators in FFP may modulate the functioning of monocytes. Interestingly, this effect appears to be partially influenced by the age of donors.

Myocardial connexin-43 and N-Cadherin decrease during vanadium inhalation.

Particulate matter air pollution has considerably increased during the last decades; vanadium is a transition element adhered to this particulate matter, and the combustion of fossil fuels is the main source in the atmosphere. It has been reported that air pollution and specifically vanadium exposure increases the probability of suffering arrhythmias; however the biological mechanism of such a relationship remains unknown. It has been established that a diminished presence of N-Cadherin alters the Connexin-43 arrangement, and the consequent altered presence of these proteins predisposes to ventricular heart rate problems. We analyzed myocardial histology and the expression of N-Cadherin and Connexin-43 by immunohistochemistry in mouse that inhaled vanadium. Our results showed a significant and progressive reduction in both N-Cadherin and Connexin-43, as well as the presence of meganucleus; myofibrils disruption, and clumping in the exposed groups were also observed. Our findings add more information about a possible explanation for the arrythmogenic effect observed in dwellers of cities with high particulate matter atmospheric pollution.

Association of Nail Dystrophy With Accrued Damage and Capillaroscopic Abnormalities in Systemic Lupus Erythematosus.

BACKGROUND: There are some nail abnormalities described in systemic lupus erythematosus (SLE). OBJECTIVES: The aim of this study was to evaluate the association between nail dystrophy (ND) and disease activity, accrued organ damage, capillaroscopic abnormalities, autoantibodies, and some markers of endothelial cell activation in patients with SLE. METHODS: This was a cross-sectional study of SLE patients from a rheumatology clinic in a tertiary care hospital. Patients were allocated in groups, according to the presence or absence of ND. Demographics, clinical data, disease activity, accrued damage, serology, nailfold capillaroscopy characteristics, serum levels of anti-endothelial cell antibodies, and plasma levels of endothelin 1 were compared between groups. Disease activity was assessed by the Systemic Lupus Erythematosus Disease Activity Index 2000 index and accrued organ damage by the Systemic Lupus International Collaborating Clinics/American College of Rheumatology Damage Index. RESULTS: Sixty-one patients were included; 50 patients (82%) were female. Thirty-two patients (52.5%) showed ND, and 29 did not. Besides a more frequent use of cyclophosphamide (46.9% vs 20.7%; P = 0.03) in the ND group, clinical features were similar. A greater organ damage was found in patients with ND (median Systemic Lupus International Collaborating Clinics/American College of Rheumatology index = 0.5, minimum = 0, maximum = 6) than in patients without ND (0, 0, 3, respectively; P = 0.04); specifically, only the skin domain was associated with ND (P = 0.04). Onycholysis (40.6%) and longitudinal ridging (25%) were the most frequent nail changes. Nailfold capillaroscopy changes were more frequent in ND patients (40.6%) than in control subjects (13.8%) (P = 0.02). The most frequent nailfold capillaroscopy findings in the ND group were enlarged capillaries (40.6%) and microhemorrhages (12.5%). There was no association between ND and the autoantibody profile, plasma endothelin 1, or serum anti-endothelial cell antibodies. CONCLUSIONS: Nail dystrophy was associated with higher accrued organ damage and the presence of capillaroscopic abnormalities. This suggests that ND might be related to chronic microvascular damage.

Statins inhibit the proliferation and induce cell death of human papilloma virus positive and negative cervical cancer cells.

Statins, competitive inhibitors of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase, have anti-tumoral effects on multiple cancer types; however, little is known about their effect on cervical cancer. We evaluated the effect on proliferation, cell cycle, oxidative stress and cell death of three statins on CaSki, HeLa (HPV(+)) and ViBo (HPV(-)) cervical cancer cell lines. Cell proliferation was assayed by crystal violet staining, cell cycle by flow cytometry and cell death by annexin-V staining. Reactive oxygen species (ROS) production was evaluated by the oxidation of 2,7-dichlorofluorescein diacetate and nitrite concentration (an indirect measure of nitric oxide (NO) production), by the Griess reaction. Inhibition of cell proliferation by atorvastatin, fluvastatin and simvastatin was dose-dependent. ViBo cells were the most responsive. Statins did not affect the cell cycle, instead they induced cell death. The antiproliferative effect in ViBo cells was completely inhibited with mevalonate, farnesyl pyrophosphate (FPP) and geranylgeranyl pyrophosphate (GGPP) treatments. In contrast, cell proliferation of CaSki and HeLa cells was partially (33%) rescued with these intermediates. The three statins increased ROS and nitrite production, mainly in the ViBo cell line. These results suggest that statins exert anti-tumoral effects on cervical cancer through inhibition of cell proliferation and induction of cell death and oxidative stress. Statins could be an aid in the treatment of cervical cancer, especially in HPV(-) tumors.

Statin-induced inhibition of MCF-7 breast cancer cell proliferation is related to cell cycle arrest and apoptotic and necrotic cell death mediated by an enhanced oxidative stress.

Statins have antiproliferative and anti-tumoral effects in MCF-7 cells. We determined the effect of statins upon MCF-7 cell cycle, toxicity, cell death, reactive oxygen species (ROS) production and mitochondrial membrane potential. Fluvastatin, simvastatin and atorvastatin inhibited cell proliferation. Antiproliferation was associated with a decrease in the DNA synthesis and a cell cycle arrest in the G1 and G2/M phases. A loss in the mitochondrial membrane potential was observed with fluvastatin. Statins induced increase in ROS production that was associated with cell death, which was abrogated by the antioxidant NAC. Our results suggest that the cytotoxic effect observed is mediated by an oxidative stress.

Altered detection of molecules associated with leukocyte traffic in HUVECs derived from newborns with a strong family history of myocardial infarction.

Atherosclerosis is a chronic inflammatory disease. As such, recruitment of immune cells is a significant event. Tightly controlled signaling molecules regulate leukocyte adhesion and migration to the tissues. The aim of this study was to determine if human umbilical vein endothelial cells (HUVECs) derived from healthy newborns with a strong family history of myocardial infarction (FHMI) showed variations in the presence of molecules related with leukocyte traffic and migration, in comparison to control healthy newborns. For this purpose, we evaluated the labeling of sialic acid containing glycoproteins, tight junction claudins and the cytoskeleton, using lectin- and immunocytochemistry in HUVECs from individuals with and without a strong FHMI. Our results show important differences in the labeling of alpha-2,3 or alpha-2,6 sialic acid-containing glycoconjugates, a disarrangement of actin filaments secondary to the absence of cytoplasmic claudin-5 immunopositivity and an increase in the binding of FHMI HUVECs to CD3+ Jurkat cells. It is possible that these differences relate to a predisposition for early appearance of atherosclerotic lesions.

Increased expression of inflammation-related co-stimulatory molecules by HUVECs from newborns with a strong family history of myocardial infarction stimulated with TNF-alpha and oxLDL.

BACKGROUND: Recent findings indicate that atherosclerosis, a chronic inflammatory process, might start during childhood. Nevertheless, the expression of inflammation-related molecules of endothelial cell isolated from healthy neonates with a strong family history of myocardial infarction (SFHMI) has been rarely analyzed. METHODS: Human umbilical vein endothelial cells (HUVECs) from children with SFHMI were assessed for the expression of CD40 and CD40L, in the presence of TNF-alpha and oxLDL. The intracellular content of CD80, CXCL8 and tissue factor by HUVECs stimulated with a CD40 agonist monoclonal antibody as well as monocytes/lymphocyte adhesion to TNF-alpha-stimulated HUVECs was also evaluated. RESULTS: The basal expression of CD40 and CD40L was higher in SFHMI-positive HUVECs in comparison to controls. TNF-alpha and oxLDL upregulated the expression of CD40 and CD40L in SFHMI versus control HUVECs (p<0.001). The intracellular expression of CXCL8, tissue factor and CD80 was also higher than in controls, and the adhesion of lymphocyte- and monocyte-like cells augmented upon TNF-alpha stimulation. CONCLUSIONS: It is possible that the modifications observed in the SFHMI-positive HUVECs, all of them relevant to the atherosclerosis process, may lead to early inflammatory reactions, thus contributing to the premature initiation of atherosclerotic lesions in these children.

HUVECs from newborns with a strong family history of diabetes show diminished ROS synthesis in the presence of high glucose concentrations.

BACKGROUND: A family history of type 2 diabetes mellitus (DM) increases the probability to develop DM and endothelial dysfunction. The probable mechanism involves augmented reactive oxygen species (ROS) synthesis. The aim of this study was to evaluate the synthesis of ROS in human umbilical vein endothelial cells (HUVECs) obtained from healthy newborns with (experimental) and without (control) a strong family history of type 2 DM, exposed to different glucose concentrations. METHODS: HUVECs were exposed to various glucose concentrations for 24 and 48 h periods, before cell proliferation, mitochondrial activity, and mitochondrial membrane potential were determined. Intracellular ROS synthesis in the presence or absence of the mitochondrial uncoupler CCCP, cytochalasin B, or diphenyleneiodonium (DPI) was also evaluated. RESULTS: As opposed to control HUVECs, we found that experimental HUVECs exposed to 30 mmol/L glucose showed a 50% decrease in cell proliferation, a 90% reduction in mitochondrial activity, and a statistically significant inhibition of ROS synthesis in the presence of CCCP or cytochalasin B; DPI had no effect. CONCLUSIONS: Our results suggest that mitochondria and NAD(P)H-oxidase from HUVECs obtained from healthy newborns with a family history of DM have an innate deficient response to high glucose concentrations.

Direct cardiac injection of G-CSF mobilized bone-marrow stem-cells improves ventricular function in old myocardial infarction.

Autologous transplant of bone marrow stem cells (BMSC), although extremely useful after acute myocardial events, has not been evaluated in patients with old (>one-year-old) myocardial infarction. Our aim was to determine if CD34(+)-enriched peripheral-blood cells, obtained by apheresis, injected directly into the severely damaged myocardium of five patients with old myocardial infarction could restore depressed myocardial function. We found that 28 weeks after revascularization and peri-infarction injection of the enriched CD34(+) peripheral mononuclear cells, ventricular hemodynamic parameters that included left ventricular ejection fraction, left ventricular diastolic volume, ventricular systolic volume and left ventricular diastolic diameter approximated normal values and there was no restenosis; two patients have been followed for >52 weeks and their parameters are within normal values. In conclusion, intramyocardial injection of easily obtained CD34(+) enriched peripheral blood cells represent an encouraging procedure for patients with severely scarred and dysfunctional myocardium.

Perspectivas de las células tallo en infartos del miocardio / Stem cell perspectives in myocardial infarctions

Myocardial infarction is the leading cause of congestive heart failure and death in industrializated countries. The cellular cardiomyoplasty has emerged as an alternative treatment in the regeneration of infarted myocardial tissue. In animals' models, differents cellular lines such as cardiomyocites, sheletal myoblast, embryonic stem cells and adult mesenchymal stem cells has been used, resulting in an improvement in ventricular function and decrease in amount of infarted tissue. The first three cells line have disvantages as they are allogenics and are difficult to obtain. The adult mesenchymal stem cells are autologous and can be obtained throught the aspiration of bone marrow or from peripherical circulation, prior to stimulating with cytokines (G-CSF). The implantation in humans with recent and old myocardial infarction have shown improvements similar to those shown in animal models. These findings encourage the continued investigation in the mechanism of cellular differentiation and implantation metods in infarted myocardial tissue.

El infarto del miocardio es la principal causa de falla cardiaca y muerte en países industrializados. A la fecha, la cardiomioplastia celular ha emergido como una alternativa en la regeneración de infartos miocárdicos. En modelos animales se han utilizado diferentes líneas celulares como cardiomiocitos fetales, mioblastos de músculo esquelético, células tallo embrionarias y células tallo mesenquimales del adulto, con mejoría en la función ventricular y disminución del área de tejido infartado. Las tres primeras líneas celulares tienen desventajas porque son alogénicas y difíciles de obtener. Las células tallo mesenquimales del adulto son autólogas y se pueden obtener de aspirados de médula ósea o de la circulación periférica previa estimulación con citocinas (G-CSF). La implantación de estas células en seres humanos con infartos del miocardio recientes y antiguos han mostrado mejorías similares a los reportes con modelos animales. Estos hallazgos alientan a continuar la investigación clínica y básica en busca de los mecanismos de diferenciación celular y selección de vías de implantación, en tejido miocárdico infartado.

[Stem cell perspectives in myocardial infarctions]. / Perspectivas de las células tallo en infartos del miocardio.

Myocardial infarction is the leading cause of congestive heart failure and death in industrializated countries. The cellular cardiomyoplasty has emerged as an alternative treatment in the regeneration of infarted myocardial tissue. In animals' models, different cellular lines such as cardiomyocites, skeletal myoblasts, embryonic stem cells and adult mesenchymal stem cells have been used, resulting in an improvement in ventricular function and decrease in amount of infarcted tissue. The first three cells lines have disvantages as they are allogenics and are difficult to obtain. The adult mesenchymal stem cells are autologous and can be obtained throught the aspiration of bone marrow or from peripherical circulation, after stimulating with cytokines (G-CSF). The implantation in humans with recent and old myocardial infarction have shown improvements similar to those shown in animal models. These findings encourage the continued investigation in the mechanism of cellular differentiation and implantation methods in infarcted myocardial tissue.