Regulation of Cardiomyocyte Proliferation by microRNAs and Small Molecules

Understanding the molecular mechanisms regulating cardiac cell proliferation during the embryonic, fetal and adult life holds a paramount importance in view of developing innovative strategies aimed at inducing myocardial regeneration after cardiac damage. Previous high throughput screening studies in our laboratory identified a series of microRNAs able to trigger cardiomyocyte proliferation and stimulate cardiac regeneration after myocardial infarction. In the first part of this project, we investigated the mechanism of action of the top ten most effective of these miRNAs, revealing an involvement of the Hippo-YAP pathway in their action. We found that all the investigated miRNAs activated YAP-mediated transcription, nuclear localization of active YAP and increased expression of YAP responsive genes. Of notice, miR-199a-3p, one of the most effective miRNAs exerted its direct effect on two mRNA targets impinging on the Hippo pathway, the inhibitory kinase TAOK1 and the E3 ubiquitin ligase, β−TrCP. Most of the miRNAs inducing proliferation (including miR-199a-3p) also modulated the dynamics of the actin cytoskeleton in the treated cardiomyocytes, which displayed a rounded shape and gross bundles of actin fibers at the cytoplasm periphery. Consistent with these observations, we found that the Cofilin2 mRNA was a direct target of four of the investigated miRNAs and that downregulation of Cofilin2 itself was sufficient to promote cardiomyocyte proliferation, activate nuclear translocation of YAP and stimulate transcription of TEAD-responsive genes. The second part of the project was aimed at identifying small molecules exerting a mitogenic effect on neonatal cardiomyocytes through an unbiased high-throughput screening (HTS) of a library of 780 FDA-registered drugs. The neuroactive alkaloid harmine was identified as the most powerful molecule at inducing cardiomyocyte proliferation in vitro and heart regeneration after myocardial infarction in vivo. Harmine exerted its activity through the inhibition of the dual specificity phosphorylation-regulated tyrosine kinase, Dyrk1a and, again, the activation of YAP nuclear translocation. Collectively, these results identify both YAP activation and actin cytoskeleton remodelling as major determinants of cardiomyocyte proliferation and establish the molecular basis for the development of pharmacological therapies to promote heart regeneration through the stimulation of the endogenous capacity of cardiomyocytes to proliferate

Prehistoric psychotropic consumption in Andean Chilean mummies

Hallucinogenic plants are often regarded as the main source of psychoactive drugs in antiquity to reach deep altered states of consciousness^1,2^. Many researchers believe this was particularly true during the Tiwanaku empire expansion, circa (500-1000 A.D.), along the Atacama Desert of Chile. Highly decorated snuffing tablets and tubes are often found as grave goods during this period^3,4,5,6,7,8^. Until now the type of drugs consumed in this paraphernalia has been unclear. From the modern city of Arica, naturally mummified human bodies with abundant hair provided a unique opportunity to test for hallucinogenic plants consumed in Andean prehistory. Analysis by gas chromatography and mass spectrometry demonstrated the presence of harmine. The Banisteriopsis vine, commonly called Ayahuasca, was the probable source. This is the first confirmed evidence of psychoactive plant consumption in pre-Hispanic Andean populations along the Atacama coastal region. Of the 32 mummy hair samples analyzed 3 males tested positive for harmine. This alkaloid aids in the catalysis and synergic effects of powerful hallucinogenic drugs. The consumption of harmine was likely related to medicinal practices and not exclusively ingested by shamans. Another important aspect of this evidence is that Banisteriopsis is an Amazon plant. It does not grow in the Atacama coastal region. Thus, our findings reveal extensive plant trade networks in antiquity between the coast, desert, highlands, and Amazon basin. The excellent preservation of human organic specimens, the use of gas chromatography and mass spectrometry allowed us to map and demonstrate the consumption of psychoactive compound plants in Andean prehistory. In addition, our findings open the door for future studies to debate the consumption and social role of ancient psychoactive and hallucinogenic plants

Chemical and photochemical properties of chloroharmine derivatives in aqueous solutions

Thermal and photochemical stability (ΦR), room temperature UV-vis absorption and fluorescence spectra, fluorescence quantum yields (ΦF) and lifetimes (τF), quantum yields of hydrogen peroxide (ΦH2O2) and singlet oxygen (ΦΔ) production, and triplet lifetimes (τT) have been obtained for the neutral and protonated forms of 6-chloroharmine, 8-chloroharmine and 6,8-dichloroharmine, in aqueous media. When it was possible, the effect of pH and oxygen concentration was evaluated. The nature of electronic transitions of protonated and neutral species of the three investigated chloroharmines was established using Time-Dependent Density Functional Theory (TD-DFT) calculations. The impact of all the foregoing observations on the biological role of the studied compounds is discussed.Fil: Rasse Suriani, Federico Ariel Osvaldo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); ArgentinaFil: Denofrio, Maria Paula. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); ArgentinaFil: Yañuk, Juan Gabriel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); ArgentinaFil: Gonzalez, Maria Micaela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); ArgentinaFil: Wolcan, Ezequiel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Fisicoquímicas Teóricas y Aplicadas. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Investigaciones Fisicoquímicas Teóricas y Aplicadas; ArgentinaFil: Seifermann, Marco. University of Mainz; AlemaniaFil: Erra Balsells, Rosa. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Centro de Investigaciones en Hidratos de Carbono. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Centro de Investigaciones en Hidratos de Carbono; ArgentinaFil: Cabrerizo, Franco Martín. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); Argentin

UVA photoactivation of harmol enhances its antifungal activity against the phytopathogens Penicillium digitatum and Botrytis cinerea

Phytopathogenic fungi responsible for post-harvest diseases on fruit and vegetables cause important economic losses. We have previously reported that harmol (1-methyl-9H-pyrido[3,4-b]indol-7-ol) is active against the causal agents of green and gray molds Penicillium digitatum and Botrytis cinerea, respectively. Here, antifungal activity of harmol was characterized in terms of pH dependency and conidial targets; also photodynamic effects of UVA irradiation on the antimicrobial action were evaluated. Harmol was able to inhibit the growth of both post-harvest fungal disease agents only in acidic conditions (pH 5), when it was found in its protonated form. Conidia treated with harmol exhibited membrane integrity loss, cell wall disruption, and cytoplasm disorganization. All these deleterious effects were more evident for B. cinerea in comparison to P. digitatum. When conidial suspensions were irradiated with UVA in the presence of harmol, antimicrobial activity against both pathogens was enhanced, compared to non-irradiated conditions. B. cinerea exhibited a high intracellular production of reactive oxygen species (ROS) when was incubated with harmol in irradiated and non-irradiated treatments. P. digitatum showed a significant increase in ROS accumulation only when treated with photoexcited harmol. The present work contributes to unravel the antifungal activity of harmol and its photoexcited counterpart against phytopathogenic conidia, focusing on ROS accumulation which could account for damage on different cellular targets.Fil: Olmedo, Gabriela María. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Instituto Superior de Investigaciones Biológicas. Universidad Nacional de Tucumán. Instituto Superior de Investigaciones Biológicas; ArgentinaFil: Cerioni, Luciana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Instituto Superior de Investigaciones Biológicas. Universidad Nacional de Tucumán. Instituto Superior de Investigaciones Biológicas; ArgentinaFil: Gonzalez, Maria Elisa. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); ArgentinaFil: Cabrerizo, Franco Martín. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); ArgentinaFil: Volentini, Sabrina Inès. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Instituto Superior de Investigaciones Biológicas. Universidad Nacional de Tucumán. Instituto Superior de Investigaciones Biológicas; ArgentinaFil: Rapisarda, Viviana Andrea. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Instituto Superior de Investigaciones Biológicas. Universidad Nacional de Tucumán. Instituto Superior de Investigaciones Biológicas; Argentin

Administration of Harmine and Imipramine Alters Creatine Kinase and Mitochondrial Respiratory Chain Activities in the Rat Brain

The present study evaluated mitochondrial respiratory chain and creatine kinase activities after administration of harmine (5, 10, and 15 mg/kg) and imipramine (10, 20, and 30 mg/kg) in rat brain. After acute treatment occurred an increase of creatine kinase in the prefrontal with imipramine (20 and 30 mg/kg) and harmine in all doses, in the striatum with imipramine (20 and 30 mg/kg) and harmine (5 and 10 mg/kg); harmine (15 mg/kg) decreased creatine kinase. In the chronic treatment occurred an increase of creatine kinase with imipramine (20 mg/kg), harmine (5 mg/kg) in the prefrontal with imipramine (20 and 30 mg/kg) and harmine (5 and 10 mg/kg) in the striatum. In the acute treatment, the complex I increased in the prefrontal with harmine (15 mg/kg) and in the striatum with harmine (10 mg/kg); the complex II decreased with imipramine (20 and 30 mg/kg) in the striatum; the complex IV increased with imipramine (30 mg/kg) in the striatum. In the chronic treatment, the complex I increased with harmine (5 mg/kg) in the prefrontal; the complex II increased with imipramine (20 mg/kg) in the prefrontal; the complex IV increased with harmine (5 mg/kg) in the striatum. Finally, these findings further support the hypothesis that harmine and imipramine could be involved in mitochondrial function

1-hydroxymethyl Harmine-TGFβSF Inhibitor: Inovasi Terapi Diabetes Melitus Terbaru Melalui Inisiasi Proses Regenerasi Sel β Pankreas pada Penderita DM Tipe 1 dan 2

Background: Type 1 and 2 diabetes mellitus (DM) is a chronic metabolic disease most commonly affects millions of people worldwide. Despite the differences in pathogenesis, both share one thing in common - that is the drastic depletion in the number of pancreatic β cells. Unfortunately, physiological proliferation of β cells has come to a halt starting from the first year of neonatal. To overcome this problem, researchers have been searching for molecules with the ability to induce β cells proliferation. Upon extensive screening, only harmine was proven to be the most potent β cells proliferation inducer. Furthermore, combination of harmine with TGFβSF inhibitor was found to boost harmine’s effectivity even more. Another development was also made to improve harmine’s selectivity by incorporating 1-hydroxymethyl group. Objective: Evaluate the potency of 1-hydroxymethyl harmine-TGFβSF inhibitor as a novel therapy for DM. Method: A systematic literature study was conducted with the database from Pubmed, Google Scholar, ScienceDirect, and Proquest for articles published within 2015-2019. Discussion: This literature review yields result that harmine-TGFβSF inhibitor is proven to induce β cells proliferation up to 18%/day or equal to 18 times the normal cell proliferation rate during embryogenesis. Moreover, incorporating 1-hydroxymethyl group into harmine is proven not only to improve selectivity but also lessen the toxicity, making 1-hydroxymethyl harmine safe as a novel therapy for diabetes. Conclusion: 1-hydroxymethyl harmine-TGFβSF inhibitor display promising potential as a novel therapy for all type of diabetes patients. Keywords: diabetes mellitus, harmine, TGFβSF inhibitor, β cell proliferation   Latar Belakang: Diabetes Melitus (DM) tipe 1 maupun tipe 2 merupakan penyakit metabolik kronis yang paling banyak ditemukan di seluruh dunia. Walaupun memiliki proses patogenesis yang berbeda, namun kedua tipe DM ini ternyata memiliki kesamaan, yaitu terjadinya penurunan kuantitas sel β pankreas. Sayangnya, kemampuan regenerasi sel β pankreas manusia telah terhenti semenjak tahun pertama masa neonatal. Untuk menangani permasalahan tersebut, para peneliti menemukan sebuah molekul bernama harmine yang terbukti efektif menginisiasi proses regenerasi sel β pankreas. Selanjutnya, untuk meningkatkan efektifitas dari harmine agar lebih baik lagi, peneliti kemudian mengkombinasikan harmine dengan TGFβSF inhibitor. Sedangkan, untuk meningkatkan selektivitas dari harmine, peneliti menambahkan gugus 1-hidroksimetil pada molekul tersebut. Tujuan: Evaluasi potensi 1-hydroxymethyl harmine-TGFβSF inhibitor sebagai terapi utama bagi semua penderita DM. Metode: Penelitian dilakukan dengan melakukan tinjauan pustaka dari beberapa database jurnal, yakni PubMed, Google Scholar, ScienceDirect dan ProQuest dengan kriteria literatur dipublikasikan dalam kurun waktu 2015-2019. Pembahasan: Studi literatur ini menunjukan bahwa harmine-TGFβSF inhibitor telah terbukti mampu meningkatkan proliferasi sel β pankreas manusia hingga mencapai 18%/hari atau setara dengan 18 kali kecepatan embriogenesis pada sel normal. Selain itu, penambahan gugus 1-hidroksimetil pada harmine juga telah terbukti tidak hanya mampu meningkatkan selektivitas dari molekul tersebut, tetapi juga mampu menurunkan efek toksisitasnya, sehingga aman digunakan sebagai terapi anti-diabetes terbaru. Kesimpulan: 1-hydroxymethyl harmine-TGFβSF inhibitor memiliki potensi yang menjanjikan untuk menjadi terapi baru bagi semua tipe penderita DM. Kata Kunci: diabetes mellitus, harmine, proliferasi sel β, TGFβSF inhibito

Harmine activates intrinsic and extrinsic pathways of apoptosis in B16F-10 melanoma

<p>Abstract</p> <p>Background</p> <p>Harmine is a beta-carboline alkaloid from the plant <it>Peganum harmala</it>. Previous studies found that harmine inhibited metastasis of B16F-10 melanoma cells. This study aims to elucidate the role of harmine in apoptosis of B16F-10 cells.</p> <p>Methods</p> <p>B16F-10 melanoma cells were treated in the presence and absence of harmine <it>in vitro</it>. Morphological changes, cell cycle and expression of various pro and anti- apoptotic genes were analyzed for the study of apoptosis.</p> <p>Results</p> <p>Morphological observation and DNA laddering assay showed that harmine treated cells displayed marked apoptotic characteristics, such as nuclear fragmentation, appearance of apoptotic bodies and DNA laddering fragment. TUNEL assay and flow cytometric analysis also confirmed apoptosis. Furthermore, RT-PCR analysis showed that harmine induced apoptosis in B16F-10 melanoma cells by up-regulating Bax and activating Caspase-3, 9 and p53 and down-regulating Bcl-2. Harmine also up-regulated Caspase-8 and Bid, indicating that harmine affected both extrinsic and intrinsic pathways of apoptosis. This study also showed inhibitory effects of harmine on some transcription factors and pro- inflammatory cytokines that protect cell from apoptosis.</p> <p>Conclusion</p> <p>Harmine activates both intrinsic and extrinsic pathways of apoptosis and regulates some transcription factors and pro-inflammatory cytokines.</p

The cell cycle regulatory DREAM complex is disrupted by high expression of oncogenic B-Myb.

Overexpression of the oncogene MYBL2 (B-Myb) is associated with increased cell proliferation and serves as a marker of poor prognosis in cancer. However, the mechanism by which B-Myb alters the cell cycle is not fully understood. In proliferating cells, B-Myb interacts with the MuvB core complex including LIN9, LIN37, LIN52, RBBP4, and LIN54, forming the MMB (Myb-MuvB) complex, and promotes transcription of genes required for mitosis. Alternatively, the MuvB core interacts with Rb-like protein p130 and E2F4-DP1 to form the DREAM complex that mediates global repression of cell cycle genes in G0/G1, including a subset of MMB target genes. Here, we show that overexpression of B-Myb disrupts the DREAM complex in human cells, and this activity depends on the intact MuvB-binding domain in B-Myb. Furthermore, we found that B-Myb regulates the protein expression levels of the MuvB core subunit LIN52, a key adapter for assembly of both the DREAM and MMB complexes, by a mechanism that requires S28 phosphorylation site in LIN52. Given that high expression of B-Myb correlates with global loss of repression of DREAM target genes in breast and ovarian cancer, our findings offer mechanistic insights for aggressiveness of cancers with MYBL2 amplification, and establish the rationale for targeting B-Myb to restore cell cycle control

Harmine Ameliorates Cognitive Impairment by Inhibiting NLRP3 Inflammasome Activation and Enhancing the BDNF/TrkB Signaling Pathway in STZ-Induced Diabetic Rats

Diabetes mellitus (DM) is considered a risk factor for cognitive dysfunction. Harmine not only effectively improves the symptoms of DM but also provides neuroprotective effects in central nervous system diseases. However, whether harmine has an effect on diabetes-induced cognitive dysfunction and the underlying mechanisms remain unknown. In this study, the learning and memory abilities of rats were evaluated by the Morris water maze test. Changes in the nucleotide-binding oligomerization domain-containing protein (NOD)-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome and brain-derived neurotrophic factor (BDNF)/TrkB signaling pathway were determined in both streptozotocin (STZ)-induced diabetic rats and high glucose (HG)-treated SH-SY5Y cells by western blotting and histochemistry. Herein, we found that harmine administration significantly ameliorated learning and memory impairment in diabetic rats. Further study showed that harmine inhibited NLRP3 inflammasome activation, as demonstrated by reduced NLRP3, ASC, cleaved caspase-1, IL-1β, and IL-18 levels, in the cortex of harmine-treated rats with DM. Harmine was observed to have similar beneficial effects in HG-treated neuronal cells. Moreover, we found that harmine treatment enhanced BDNF and phosphorylated TrkB levels in both the cortex of STZ-induced diabetic rats and HG-treated cells. These data indicate that harmine mitigates cognitive impairment by inhibiting NLRP3 inflammasome activation and enhancing the BDNF/TrkB signaling pathway. Thus, our findings suggest that harmine is a potential therapeutic drug for diabetes-induced cognitive dysfunction

Selective inhibition of toxic cyanobacteria by β-carboline-containing bacterium Bacillus flexus isolated from Saudi freshwaters

AbstractA bacterial strain SSZ01 isolated from a eutrophic lake in Saudi Arabia dominated by cyanobacterial blooms, showed an antialgal activity against cyanobacteria species. Based on the analysis of the 16S rDNA gene sequence, the isolated strain (SSZ01) most likely belonged to the genus Bacillus with a 99% similarity to Bacillus flexus strain EMGA5. The thin layer chromatography (TLC) analysis of the ethyl acetate extract of this bacterium revealed that this strain can produce harmine and norharmane compared to different β-carboline analog standards. Harmine and norharmane were also detected in considerable amounts in bacterial growth medium, indicating a potential excretion of these compounds into the aquatic environment. The crude extract of Bacillus flexus as well as pure materials of harmine and norharmane inhibited the growth of tested species of cyanobacteria. However, the bacterial crude extract has a higher toxicity against tested species of cyanobacteria than harmine and norharmane. In addition, harmine was more toxic to cyanobacteria than norharmane. On the other hand, neither pure compounds of harmine and norharmane nor crude bacterial extract showed any antialgal activity against tested species of green algae. The results of the present study suggest that B. flexus SSZ01 or its crude extract containing harmine and norharmane could be a candidate for the selective control of cyanobacterial blooms without affecting other algal species