bims-tubesc 2021-05-09 papers

bims-tubesc

Biomed News

on Molecular mechanisms in tuberous sclerosis

Issue of 2021–05–09
thirteen papers selected by

Conditional knockout of Tsc1 in RORγt-expressing cells induces brain damage and early death in mice.
Diabetes in Individuals With Tuberous Sclerosis Complex Treated With mTOR Inhibitors.
MicroRNA-34a activation in tuberous sclerosis complex during early brain development may lead to impaired corticogenesis.
Increased branched-chain amino acids at baseline and hours before a spontaneous seizure in the human epileptic brain.
Tuberous sclerosis complex: The critical role of the interventional radiologist in management.
TORC1 regulates the transcriptional response to glucose and developmental cycle via the Tap42-Sit4-Rrd1/2 pathway in Saccharomyces cerevisiae.
Speech Analysis Using Artificial Intelligence as a Peri-Operative Evaluation: A Case Report of a Patient with Temporal Lobe Epilepsy Secondary to Tuberous Sclerosis Complex Who Underwent Epilepsy Surgery.
Giant bilateral angiomyolipoma of the kidney.
mTOR signaling regulates gastric epithelial progenitor homeostasis and gastric tumorigenesis via MEK1-ERKs and BMP-Smad1 pathways.
Low-Grade Oncocytic Tumor of Kidney (CK7-Positive, CD117-Negative): Incidence in a Single Institutional Experience with Clinicopathological and Molecular Characteristics.
Genetic heterogeneity of disorders with overgrowth and intellectual disability: Experience from a center in North India.
Increasing mTORC1 Pathway Activity or Methionine Supplementation during Pregnancy Reverses the Negative Effect of Maternal Malnutrition on the Developing Kidney.
CHIR99021 Augmented the Function of Late Endothelial Progenitor Cells by Preventing Replicative Senescence.

J Neuroinflammation. 2021 May 06. 18(1): 107

Conditional knockout of Tsc1 in RORγt-expressing cells induces brain damage and early death in mice.

Yafei Deng, Qinglan Yang, Yao Yang, Yana Li, Hongyan Peng, Shuting Wu, Shuju Zhang, Baige Yao, Shuhui Li, Yuan Gao, Xiaohui Li, Liping Li, Youcai Deng.

   BACKGROUND: Tuberous sclerosis complex 1 (Tsc1) is known to regulate the development and function of various cell types, and RORγt is a critical transcription factor in the immune system. However, whether Tsc1 participates in regulating RORγt-expressing cells remains unknown.
METHODS: We generated a mouse model in which Tsc1 was conditionally deleted from RORγt-expressing cells (Tsc1RORγt) to study the role of RORγt-expressing cells with Tsc1 deficiency in brain homeostasis.
RESULTS: Type 3 innate lymphoid cells (ILC3s) in Tsc1RORγt mice displayed normal development and function, and the mice showed normal Th17 cell differentiation. However, Tsc1RORγt mice exhibited spontaneous tonic-clonic seizures and died between 4 and 6 weeks after birth. At the age of 4 weeks, mice in which Tsc1 was specifically knocked out in RORγt-expressing cells had cortical neuron defects and hippocampal structural abnormalities. Notably, over-activation of neurons and astrogliosis were observed in the cortex and hippocampus of Tsc1RORγt mice. Moreover, expression of the γ-amino butyric acid (GABA) receptor in the brains of Tsc1RORγt mice was decreased, and GABA supplementation prolonged the lifespan of the mice to some extent. Further experiments revealed the presence of a group of rare RORγt-expressing cells with high metabolic activity in the mouse brain.
CONCLUSIONS: Our study verifies the critical role of previously unnoticed RORγt-expressing cells in the brain and demonstrates that the Tsc1 signaling pathway in RORγt-expressing cells is important for maintaining brain homeostasis.

Keywords:  GABA; RORγt-expressing cells; Seizure; Tsc1

DOI:  https://doi.org/10.1186/s12974-021-02153-8
Pediatr Neurol. 2021 Apr 02. pii: S0887-8994(21)00053-9. [Epub ahead of print]120 7-10

Diabetes in Individuals With Tuberous Sclerosis Complex Treated With mTOR Inhibitors.

Karen Agricola, Gabrielle Stires, Darcy A Krueger, Jamie K Capal, David N Franz, David M Ritter.

   BACKGROUND: Tuberous sclerosis complex (TSC) is a genetic disorder that is manifested in multiple body systems. A mammalian target of rapamycin (mTOR) inhibitor (mTORi), either everolimus or sirolimus, is now routinely prescribed for multiple clinical manifestations of TSC, including subependymal giant cell astrocytoma and epilepsy. These medications are generally well tolerated. Side effects previously identified in well-designed clinical trials tend to be mild and readily manageable. Regulatory approvals for the treatment of TSC have expanded the use of everolimus and sirolimus clinically, enlarging clinician experience and enabling identification of potential treatment-related effects that are rarer than could be identified or recognized in previous clinical trials.
METHODS: The medical records of clinical patients from our TSC center who were treated with an mTORi and later developed diabetes mellitus (DM) were analyzed and compared with those who were not treated with an mTORi. Eight individuals received detailed analysis, including laboratory results, concomitant medications, and body mass indices.
RESULTS: Among the 1576 individuals with TSC, 4% taking an mTORi developed diabetes compared with 0.6% of those not on mTORi, showing a significant interaction between DM and mTORi (chi-square = 18.1, P < 0.001). Details of eight patients who developed DM were presented.
CONCLUSIONS: The long-term use of mTORi agents in TSC may contribute to a risk of diabetes. Early detection can be critical in management. Additional studies are need to further investigate a causal relationship, but clinicians should be aware of this possible association when initiating and monitoring ongoing treatment.

Keywords:  Adverse events; Diabetes mellitus; Tuberous sclerosis; mTOR inhibitors

DOI:  https://doi.org/10.1016/j.pediatrneurol.2021.03.007
Neuropathol Appl Neurobiol. 2021 May 03.

MicroRNA-34a activation in tuberous sclerosis complex during early brain development may lead to impaired corticogenesis.

A Korotkov, N S Sim, M J Luinenburg, J J Anink, J van Scheppingen, T S Zimmer, A Bongaarts, D W M Broekaart, C Mijnsbergen, F E Jansen, W Van Hecke, W G M Spliet, P C van Rijen, M Feucht, J A Hainfellner, P Krsek, J Zamecnik, P B Crino, K Kotulska, L Lagae, A C Jansen, D J Kwiatkowski, S Jozwiak, P Curatolo, A Mühlebner, J H Lee, J D Mills, E A van Vliet, E Aronica.

   AIMS: Tuberous sclerosis complex (TSC) is a genetic disorder associated with dysregulation of the mechanistic target of rapamycin complex 1 (mTORC1) signalling pathway. Neurodevelopmental disorders, frequently present in TSC, are linked to cortical tubers in the brain. We previously reported microRNA-34a (miR-34a) among the most up-regulated miRs in tubers. Here, we characterized miR-34a expression in tubers with the focus on the early brain development and assessed the regulation of mTORC1 pathway and corticogenesis by miR-34a.
METHODS: We analysed the expression of miR-34a in resected cortical tubers (n = 37) compared to autopsy-derived control tissue (n = 27). The effect of miR-34a overexpression on corticogenesis was assessed in mice at E18. The regulation of the mTORC1 pathway and the expression of the bioinformatically predicted target genes were assessed in primary astrocyte cultures from 3 patients with TSC and in SH-SY5Y cells following miR-34a transfection.
RESULTS: The peak of miR-34a overexpression in tubers was observed during infancy, concomitant with the presence of pathological markers, particularly in giant cells and dysmorphic neurons. MiR-34a was also strongly expressed in fetal TSC cortex. Overexpression of miR-34a in mouse embryos decreased the percentage of cells migrated to the cortical plate. The transfection of miR-34a mimic in TSC astrocytes negatively regulated mTORC1 and decreased the expression of the target genes RAS related (RRAS) and NOTCH1.
CONCLUSIONS: MiR-34a is most highly overexpressed in tubers during fetal and early postnatal brain development. MiR-34a can negatively regulate mTORC1, however, it may also contribute to abnormal corticogenesis in TSC.

Keywords:  TSC; mechanistic target of rapamycin; miRNA; migration; neurodevelopmental disorder

DOI:  https://doi.org/10.1111/nan.12717
Epilepsia. 2021 May 05.

Increased branched-chain amino acids at baseline and hours before a spontaneous seizure in the human epileptic brain.

Caroline Ong, Eyiyemisi C Damisah, Shaun E Gruenbaum, Roni Dhaher, Yanhong Deng, Mani Ratnesh S Sandhu, Hitten P Zaveri, Dennis D Spencer, Tore Eid.

  The objective of this study was to monitor the extracellular brain chemistry dynamics at baseline and in relation to spontaneous seizures in human patients with refractory epilepsy. Thirty patients with drug-resistant focal epilepsy underwent intracranial electroencephalography and concurrent brain microdialysis for up to 8 continuous days. Extracellular brain glutamate, glutamine, and the branched-chain amino acids (BCAAs) valine, leucine, and isoleucine were quantified in the dialysis samples by liquid chromatography-tandem mass spectrometry. Extracellular BCAAs and glutamate were chronically elevated at baseline by approximately 1.5-3-fold in brain regions of seizure onset and propagation versus regions not involved by seizures. Moreover, isoleucine increased significantly above baseline as early as 3 h before a spontaneous seizure. BCAAs play important roles in glutamatergic neurotransmission, mitochondrial function, neurodegeneration, and mammalian target of rapamycin signaling. Because all of these processes have been implicated in epilepsy, the results suggest a novel role of BCAAs in the pathogenesis of spontaneous seizures.

Keywords:  ictogenesis; metabolism; microdialysis; neurochemistry; seizure prediction

DOI:  https://doi.org/10.1111/epi.16920
SA J Radiol. 2021 ;25(1): 2034

Tuberous sclerosis complex: The critical role of the interventional radiologist in management.

Puneet Garg, Anuradha Sharma, Heena Rajani, Apratim R Choudhary, Rajkumar Meena.

  Tuberous sclerosis complex (TSC) is an autosomal dominant neurocutaneous syndrome that is characterised by hamartomas in multiple organs, the characteristic imaging features of which are illustrated in this case report. Angiomyolipoma (AML) is the most common renal manifestation of TSC, which may present with life-threatening haemorrhage at the time of diagnosis. Interventional management with selective renal embolisation is currently the treatment of choice for the safe and effective management of ruptured renal AML.

Keywords:  angiomyolipoma; hamartomas; renal haemorrhage; ruptured angiomyolipoma; selective renal embolisation; tuberous sclerosis complex

DOI:  https://doi.org/10.4102/sajr.v25i1.2034
BMC Biol. 2021 May 06. 19(1): 95

TORC1 regulates the transcriptional response to glucose and developmental cycle via the Tap42-Sit4-Rrd1/2 pathway in Saccharomyces cerevisiae.

Mohammad Alfatah, Jin Huei Wong, Vidhya Gomathi Krishnan, Yong Cheow Lee, Quan Feng Sin, Corinna Jie Hui Goh, Kiat Whye Kong, Wei Ting Lee, Jacqueline Lewis, Shawn Hoon, Prakash Arumugam.

   BACKGROUND: Target of Rapamycin Complex 1 (TORC1) is a highly conserved eukaryotic protein complex that couples the presence of growth factors and nutrients in the environment with cellular proliferation. TORC1 is primarily implicated in linking amino acid levels with cellular growth in yeast and mammals. Although glucose deprivation has been shown to cause TORC1 inactivation in yeast, the precise role of TORC1 in glucose signaling and the underlying mechanisms remain unclear.
RESULTS: We demonstrate that the presence of glucose in the growth medium is both necessary and sufficient for TORC1 activation. TORC1 activity increases upon addition of glucose to yeast cells growing in a non-fermentable carbon source. Conversely, shifting yeast cells from glucose to a non-fermentable carbon source reduces TORC1 activity. Analysis of transcriptomic data revealed that glucose and TORC1 co-regulate about 27% (1668/6004) of yeast genes. We demonstrate that TORC1 orchestrates the expression of glucose-responsive genes mainly via the Tap42-Sit4-Rrd1/2 pathway. To confirm TORC1's function in glucose signaling, we tested its role in spore germination, a glucose-dependent developmental state transition in yeast. TORC1 regulates the glucose-responsive genes during spore germination and inhibition of TORC1 blocks spore germination.
CONCLUSIONS: Our studies indicate that a regulatory loop that involves activation of TORC1 by glucose and regulation of glucose-responsive genes by TORC1, mediates nutritional control of growth and development in yeast.

Keywords:  Spore germination; TORC1; Tap42/Sit4/Rrd1-2 module; Transcriptional response to glucose

DOI:  https://doi.org/10.1186/s12915-021-01030-3
Brain Sci. 2021 Apr 29. pii: 568. [Epub ahead of print]11(5):

Speech Analysis Using Artificial Intelligence as a Peri-Operative Evaluation: A Case Report of a Patient with Temporal Lobe Epilepsy Secondary to Tuberous Sclerosis Complex Who Underwent Epilepsy Surgery.

Keiko Niimi, Ayataka Fujimoto, Yoshinobu Kano, Yoshiro Otsuki, Hideo Enoki, Tohru Okanishi.

   BACKGROUND: Improved conversational fluency is sometimes identified postoperatively in patients with epilepsy, but improvements can be difficult to assess using tests such as the intelligence quotient (IQ) test. Evaluation of pre- and postoperative differences might be considered subjective at present because of the lack of objective criteria. Artificial intelligence (AI) could possibly be used to make the evaluations more objective. The aim of this case report is thus to analyze the speech of a young female patient with epilepsy before and after surgery.
METHOD: The speech of a nine-year-old girl with epilepsy secondary to tuberous sclerosis complex is recorded during interviews one month before and two months after surgery. The recorded speech is then manually transcribed and annotated, and subsequently automatically analyzed using AI software. IQ testing is also conducted on both occasions. The patient remains seizure-free for at least 13 months postoperatively.
RESULTS: There are decreases in total interview time and subjective case markers per second, whereas there are increases in morphemes and objective case markers per second. Postoperatively, IQ scores improve, except for the Perceptual Reasoning Index.
CONCLUSIONS: AI analysis is able to identify differences in speech before and after epilepsy surgery upon an epilepsy patient with tuberous sclerosis complex.

Keywords:  artificial intelligence; epilepsy; intelligence quotient; speech analysis; surgery

DOI:  https://doi.org/10.3390/brainsci11050568
Ann R Coll Surg Engl. 2021 May 06.

Giant bilateral angiomyolipoma of the kidney.

R Bhatia, L Bains, P Lal, G Bhagria.

  Angiomyolipoma is a benign solid renal neoplasm. A giant angiomyolipoma is more than 10cm by size, but it can grow to huge proportions. Our case appears to be the third largest angiomyolipoma and the largest among bilateral giant renal angiomyolipoma in the indexed literature. A 26-year-old man presented with large right abdominal swelling for the past three years, which was occupying his right flank and iliac region, extending beyond the midline. Computed tomography of the abdomen revealed a large well-defined mass in the right side of the abdomen, crossing the midline and measuring 35 × 20 × 12cm. The left kidney showed a similar fatty lesion of 14 × 6cm. The findings were consistent with angiomyolipoma. Further evaluation for tuberous sclerosis by magnetic resonance imaging the brain demonstrated multiple subependymal nodules. Giant renal angiomyolipoma is an uncommon tumour with bilateral giant angiomyolipoma being a rare entity. Preoperative embolisation helps in reducing size of the tumour. In case of giant and bilateral angiomyolipoma, evaluation for tuberous sclerosis should always be done.

Keywords:  Angiomyolipoma; Bilateral; Giant; Tuberous sclerosis

DOI:  https://doi.org/10.1308/rcsann.2020.7036
Cell Rep. 2021 May 04. pii: S2211-1247(21)00400-9. [Epub ahead of print]35(5): 109069

mTOR signaling regulates gastric epithelial progenitor homeostasis and gastric tumorigenesis via MEK1-ERKs and BMP-Smad1 pathways.

Ke Li, Hongguang Wu, Ao Wang, Jean Charron, Yuji Mishina, Samy L Habib, Huijuan Liu, Baojie Li.

  mTOR, the sensor of nutrients and growth factors, has important roles in tissue homeostasis and tumorigenesis. However, how mTOR controls gastric epithelial cell turnover and gastric cancer development, a leading malignancy, remains poorly understood. Here, we provide genetic evidence that mTOR activation promotes proliferation and inhibits differentiation of Lgr5+ gastric epithelial progenitors (GEPs) in gastric homeostasis and tumorigenesis. mTOR signaling increases MEK1 and Smad1 expression and enhances activation of MEK1-ERKs and BMP-Smad1 pathways, respectively, in GEPs and gastric tumors. Mek1 deletion or inhibition rescues hyperproliferation, whereas Bmpr1a ablation or inhibition rescues differentiation defects of Tsc1-/- GEPs. Tsc1 deficiency in Lgr5+ GEPs accelerates gastric tumor initiation and development, which require MEK1-ERKs for hyperplasia and BMP-Smad1 for differentiation suppression. These findings reveal how mTOR signaling controls Lgr5+ GEP homeostasis and cancerization and suggest that ERKs and Smad1 signaling can be safely targeted to substitute mTOR inhibitors in gastric cancer therapy.

Keywords:  Lgr5; MEK1; Smad1; TSC; differentiation; gastric cancer; gastric epithelial progenitor; homeostasis; mTOR; tumorigenesis

DOI:  https://doi.org/10.1016/j.celrep.2021.109069
Hum Pathol. 2021 May 04. pii: S0046-8177(21)00071-X. [Epub ahead of print]

Low-Grade Oncocytic Tumor of Kidney (CK7-Positive, CD117-Negative): Incidence in a Single Institutional Experience with Clinicopathological and Molecular Characteristics.

Oleksandr Kravtsov, Sounak Gupta, John C Cheville, William R Sukov, Ross Rowsey, Loren P Herrera-Hernandez, Christine M Lohse, Ryan Knudson, Bradley C Leibovich, Rafael E Jimenez.

   AIMS: Low-grade oncocytic tumor of the kidney (LOT) is characterized by cytoplasmic eosinophilia and a CK7-positive/CD117-negative immunophenotype. Morphologically, they exhibit overlapping features with oncocytoma and chromophobe renal cell carcinoma (chRCC). Our aim was to obtain long-term clinical follow-up data, clinicopathological and molecular characteristics, and incidence of LOT.
METHODS: Tissue microarrays were constructed from 574 tumors historically diagnosed as oncocytoma and surgically treated at Mayo Clinic between 1970 and 2012, and immunostained for CK7 and CD117. An extended immunophenotype was obtained on whole slide sections, along with FISH for CCND1 rearrangement status and chromosomal microarray for copy number status. Additionally, two cases were retrospectively identified in a set of tuberous sclerosis complex (TSC)-associated neoplasms and three more cases diagnosed on needle core biopsies were obtained during routine clinical practice.
RESULTS: Twenty-four cases of LOT were identified amongst 574 consecutive tumors diagnosed as oncocytoma and treated with partial or radical nephrectomy, corresponding to an incidence of 4.18% of tumors historically diagnosed as oncocytomas, and 0.35% of 6944 nephrectomies performed between 1970 and 2012. Overall, 29 cases of LOT were identified in three clinical settings: sporadic, TSC-associated, and end-stage renal disease (ESRD). Multifocality was seen only in the setting of TSC and ESRD. No metastases attributable to LOT were identified (median follow up 9.6 years). There were no recurrent arm level copy number changes detected by chromosomal microarray and all tested cases were negative for CCND1 rearrangement by FISH.
CONCLUSIONS: LOT is an uncommon eosinophilic renal neoplasm with an indolent prognosis that constitutes ∼ 4% of tumors historically diagnosed as oncocytoma. The morphologic, immunophenotypic, and molecular features of this neoplasm suggest it is a distinct entity of renal neoplasia.

Keywords:  Low-grade oncocytic renal tumor; chromophobe renal cell carcinoma; cytogenetics kidney tumors; oncocytic renal neoplasms; oncocytoma

DOI:  https://doi.org/10.1016/j.humpath.2021.04.013
Am J Med Genet A. 2021 May 04.

Genetic heterogeneity of disorders with overgrowth and intellectual disability: Experience from a center in North India.

Amita Moirangthem, Kausik Mandal, Deepti Saxena, Priyanka Srivastava, Poonam Singh Gambhir, Neha Agrawal, Arya Shambhavi, Sheela Nampoothiri, Shubha R Phadke.

  Overgrowth, defined as height and/or OFC ≥ +2SD, characterizes a subset of patients with syndromic intellectual disability (ID). Many of the disorders with overgrowth and ID (OGID) are rare and the full phenotypic and genotypic spectra have not been unraveled. This study was undertaken to characterize the phenotypic and genotypic profile of patients with OGID. Patients with OGID were ascertained from the cohort of patients who underwent cytogenetic microarray (CMA) and/or exome sequencing (ES) at our center over a period of 6 years. Thirty-one subjects (six females) formed the study group with ages between 3.5 months and 13 years. CMA identified pathogenic deletions in two patients. In another 11 patients, a disease causing variant was detected by ES. The spectrum of disorders encompassed aberrations in genes involved in the two main pathways associated with OGID. These were genes involved in epigenetic regulation like NSD1, NFIX, FOXP1, and those in the PI3K-AKT pathway like PTEN, AKT3, TSC2, PPP2R5D. Five novel pathogenic variants were added by this study. NSD1-related Sotos syndrome was the most common disorder, seen in five patients. A causative variant was identified in 61.5% of patients who underwent only ES compared to the low yield of 11.1% in the CMA group. The molecular etiology could be confirmed in 13 subjects with OGID giving a diagnostic yield of 42%. The major burden was formed by autosomal dominant monogenic disorders. Hence, ES maybe a better first-tier genomic test rather than CMA in OGID.

Keywords:  Sotos syndrome; developmental delay; exome sequencing; macrocephaly; overgrowth

DOI:  https://doi.org/10.1002/ajmg.a.62241
J Am Soc Nephrol. 2021 May 06. pii: ASN.2020091321. [Epub ahead of print]

Increasing mTORC1 Pathway Activity or Methionine Supplementation during Pregnancy Reverses the Negative Effect of Maternal Malnutrition on the Developing Kidney.

Yaniv Makayes, Elad Resnick, Liad Hinden, Elina Aizenshtein, Tomer Shlomi, Raphael Kopan, Morris Nechama, Oded Volovelsky.

   BACKGROUND: Low nephron number at birth is associated with a high risk of CKD in adulthood because nephrogenesis is completed in utero. Poor intrauterine environment impairs nephron endowment via an undefined molecular mechanism. A calorie-restricted diet (CRD) mouse model examined the effect of malnutrition during pregnancy on nephron progenitor cells (NPCs).
METHODS: Daily caloric intake was reduced by 30% during pregnancy. mRNA expression, the cell cycle, and metabolic activity were evaluated in sorted Six2 NPCs. The results were validated using transgenic mice, oral nutrient supplementation, and organ cultures.
RESULTS: Maternal CRD is associated with low nephron number in offspring, compromising kidney function at an older age. RNA-seq identified cell cycle regulators and the mTORC1 pathway, among other pathways, that maternal malnutrition in NPCs modifies. Metabolomics analysis of NPCs singled out the methionine pathway as crucial for NPC proliferation and maintenance. Methionine deprivation reduced NPC proliferation and lowered NPC number per tip in embryonic kidney cultures, with rescue from methionine metabolite supplementation. Importantly, in vivo, the negative effect of caloric restriction on nephrogenesis was prevented by adding methionine to the otherwise restricted diet during pregnancy or by removing one Tsc1 allele in NPCs.
CONCLUSIONS: These findings show that mTORC1 signaling and methionine metabolism are central to the cellular and metabolic effects of malnutrition during pregnancy on NPCs, contributing to nephrogenesis and later, to kidney health in adulthood.

Keywords:  intrauterine environment; kidney development; mTOR pathway; malnutrition; methionine; nephron progenitor cells; stem cell

DOI:  https://doi.org/10.1681/ASN.2020091321
Int J Mol Sci. 2021 Apr 30. pii: 4796. [Epub ahead of print]22(9):

CHIR99021 Augmented the Function of Late Endothelial Progenitor Cells by Preventing Replicative Senescence.

Vinoth Kumar Rethineswaran, Da Yeon Kim, Yeon-Ju Kim, WoongBi Jang, Seung Taek Ji, Le Thi Hong Van, Ly Thanh Truong Giang, Jong Seong Ha, Jisoo Yun, Jinsup Jung, Sang-Mo Kwon.

  Endothelial progenitor cells (EPCs) are specialized cells in circulating blood, well known for their ability to form new vascular structures. Aging and various ailments such as diabetes, atherosclerosis and cardiovascular disease make EPCs vulnerable to decreasing in number, which affects their migration, proliferation and angiogenesis. Myocardial ischemia is also linked to a reduced number of EPCs and their endothelial functional role, which hinders proper blood circulation to the myocardium. The current study shows that an aminopyrimidine derivative compound (CHIR99021) induces the inhibition of GSK-3β in cultured late EPCs. GSK-3β inhibition subsequently inhibits mTOR by blocking the phosphorylation of TSC2 and lysosomal localization of mTOR. Furthermore, suppression of GSK-3β activity considerably increased lysosomal activation and autophagy. The activation of lysosomes and autophagy by GSK-3β inhibition not only prevented replicative senescence of the late EPCs but also directed their migration, proliferation and angiogenesis. To conclude, our results demonstrate that lysosome activation and autophagy play a crucial role in blocking the replicative senescence of EPCs and in increasing their endothelial function. Thus, the findings provide an insight towards the treatment of ischemia-associated cardiovascular diseases based on the role of late EPCs.

Keywords:  CHIR99021; EPC; GSK-3β; autophagy; lysosome; mTOR; senescence

DOI:  https://doi.org/10.3390/ijms22094796