About

The Lo Sardo laboratory combines pluripotent stem cell potential and functional genomics to understand how common genetic variants among individuals, including those in non-coding portions of the genome, contribute to altering cell physiology, cell state, and fate commitment.

Research topics

• Genetics
• Biology
• Cell biology
• Endocrinology
• Immunology
• Internal medicine
• Medicine

Selected publications

• Abstract Tu0027a: The 9p21.3 coronary artery disease risk locus drives vascular smooth muscle cells to an osteochondrogenic state

  Arteriosclerosis Thrombosis and Vascular Biology · 2025-04-01

  article1st authorCorresponding

  Background: Genome-wide association studies have identified common genetic variants at ~300 human genomic loci linked to coronary artery disease (CAD) susceptibility. Among these genomic regions, the most impactful is the 9p21.3 CAD risk locus, which spans a 60 kb gene desert and encompasses ~80 SNPs in high linkage disequilibrium. Despite nearly two decades since its discovery, the role of the 9p21.3 locus in cells of the vasculature remains incompletely resolved. Methods: Vascular smooth muscle cells (VSMC) differentiation of induced pluripotent stem cells (iPSCs) from risk and non-risk donors at 9p21.3. Single-cell transcriptomic profiling, including co-embedding and comparison with publicly available human arteries single cell RNA sequencing datasets. Validation of hits on iPSC-VSMCs derived from additional human donors through immunocytochemistry, gene expression analysis and calcification staining. Migration assays of iPSC-VSMCs and primary aortic smooth muscle cells. Lentivirus-mediated overexpression of ANRIL followed by gene expression analysis. Results: iPSC-VSMCs harboring the 9p21.3 risk haplotype preferentially adopt an osteochondrogenic state and show remarkable similarity to fibrochondrocytes from human artery tissue. The transcriptional profile and functional assessment of migration and calcification capacity across iPSC-VSMCs lines from multiple donors concordantly resemble an osteochondrogenic state. Importantly, we identified numerous transcription factors driving different VSMC state trajectories. Additionally, we prioritized LIMCH1 and CRABP1 as signature genes critical for defining the risk transcriptional program. Overexpression of short ANRIL in non-risk cells is sufficient to induce the osteochondrogenic transcriptional signature. Conclusion: Our study provides new insights into the mechanism of the 9p21.3 risk locus and defines its previously undescribed role in driving a disease-prone transcriptional and functional state in VSMCs concordant with an osteochondrogenic-like state. Our data suggest that the 9p21.3 risk haplotype likely promotes arterial calcification, through altered expression of ANRIL, in a cell-type specific and cell- autonomous manner, providing insight into potential risk assessment and treatment for carriers.

  PublisherDOI

• The 9p21.3 Coronary Artery Disease Risk Locus Drives Vascular Smooth Muscle Cells to an Osteochondrogenic State

  Arteriosclerosis Thrombosis and Vascular Biology · 2025-03-27 · 8 citations

  articleOpen accessSenior author

  BACKGROUND: Genome-wide association studies have identified common genetic variants at ≈300 human genomic loci linked to coronary artery disease susceptibility. Among these genomic regions, the most impactful is the 9p21.3 coronary artery disease risk locus, which spans a 60-kb gene desert and encompasses ≈80 SNPs (single nucleotide polymorphism) in high linkage disequilibrium. Despite ≈2 decades since its discovery, the role of the 9p21.3 locus in cells of the vasculature remains incompletely resolved. METHODS: We differentiated induced pluripotent stem cells (iPSCs) from risk, nonrisk donors at 9p21.3, and isogenic knockouts into vascular smooth muscle cells (VSMCs). We performed single-cell transcriptomic profiling, including coembedding and comparison with publicly available human arterial data sets. We conducted functional characterization using migration and calcification assays and confirmed our findings on iPSC–VSMCs derived from additional donors. Finally, we used overexpression of ANRIL followed by gene expression analysis. RESULTS: We demonstrated that iPSC–VSMCs harboring the 9p21.3 risk haplotype preferentially adopt an osteochondrogenic state and show remarkable similarity to fibrochondrocytes from human artery tissue. The transcriptional profile and functional assessment of migration and calcification capacity across iPSC–VSMC lines from multiple donors concordantly resemble an osteochondrogenic state. Importantly, we identified numerous transcription factors driving different VSMC state trajectories. Additionally, we prioritized LIMCH1 and CRABP1 as signature genes critical for defining the risk transcriptional program. Finally, overexpression of a short isoform of ANRIL in 9p21.3 knockout cells was sufficient to induce the osteochondrogenic transcriptional signature. CONCLUSIONS: Our study provides new insights into the mechanism of the 9p21.3 risk locus and defines its previously undescribed role in driving a disease-prone transcriptional and functional state in VSMCs concordant with an osteochondrogenic-like state. Our data suggest that the 9p21.3 risk haplotype likely promotes arterial calcification, through altered expression of ANRIL , in a cell type–specific and cell-autonomous manner, providing insight into potential risk assessment and treatment for carriers.

  PublisherDOI

• Abstract Mo084: Stem Cell-Based Functional Genomics Unravel A Novel Vascular Smooth Muscle Cell State Induced By The 9p21 Coronary Artery Disease Risk Locus

  Circulation Research · 2024-08-02

  articleSenior author

  Background: Genome-wide association studies have identified common genetic variants (Single Nucleotide Polymorphisms – SNPs) at ~300 human genomic loci linked to coronary artery disease (CAD) susceptibility. Among these genomic regions, the most impactful is the 9p21.3 CAD risk locus, which spans a 60 kb gene desert and encompasses ~80 SNPs in high linkage disequilibrium. We have previously generated isogenic induced Pluripotent Stem Cells (iPSCs) lines from risk and non-risk donors at 9p21.3 and performed a complete deletion of the locus. By using iPSC-derived vascular smooth muscle cells (iPSC-VSMCs) we demonstrated that the risk haplotype at 9p21.3 causes altered expression of numerous genes essential for muscle function, including contraction and adhesion. Notably, deletion of the risk haplotype restores the non-risk phenotype, suggesting a gain of function effect. Here, we aimed to identify the transcriptomic signature and state trajectories of VSMCs induced by the 9p21.3 risk haplotype and validate them through functional assays and ex-vivo analysis of human coronary arteries. Methods: We have used iPSCs from individuals carrying the risk and non-risk haplotype at 9p21.3 and isogenic knockout lines. After VSMCs differentiation induction, mature VSMCs were used for single cell RNA sequencing using 10X Chromium platform and functional assays. Results: Our analysis revealed that the 9p21.3 risk haplotype prompts VSMCs to acquire a novel cellular state showing reduced plasticity and divergent from other VSMC states previously linked to CAD. We found markers of alternative lineages, including osteogenic and neuronal, coupled with altered expression of genes within other CAD loci, and functional defects. We identified a set of new molecular markers crucial to define risk-associated VSMCs and uncovered their upstream regulators. Conclusions: Our study provides insights into CAD pathogenesis driven by the 9p21.3 risk locus and identified new gene regulatory networks essential for maintaining the normal functionality of the muscle layer of the arteries. Leveraging the power of iPSCs we present novel concepts about the biology of VSMCs and shed light on the impact of the strongest CAD genetic risk factor in preserving a healthy cellular state within the vasculature.

  PublisherDOI

• Abstract 2072: The 9p21.3 Coronary Artery Disease Risk Locus Induces Cell State Transitions In Vascular Smooth Muscle Cells

  Arteriosclerosis Thrombosis and Vascular Biology · 2024-05-01

  article1st authorCorresponding

  Background: Genome-wide association studies have identified common genetic variants (Single Nucleotide Polymorphisms - SNPs) at ~300 human genomic loci linked to coronary artery disease (CAD) susceptibility. Among these genomic regions, the most impactful is the 9p21.3 CAD risk locus, which spans a 60 kb gene desert and encompasses ~80 SNPs in high linkage disequilibrium. We have previously generated isogenic induced Pluripotent Stem Cells (iPSCs) lines from risk and non-risk donors at 9p21.3 and performed a complete deletion of the locus. By using iPSC-derived vascular smooth muscle cells (iPSC-VSMCs) we demonstrated that the risk haplotype at 9p21.3 causes altered expression of numerous genes essential for muscle function, including contraction and adhesion. Notably, deletion of the risk haplotype restores the non-risk phenotype, suggesting a gain of function effect. Hypothesis: The 9p21.3 risk haplotype alters VSMCs plasticity and induces dedifferentiation and alternative lineage acquisition. Aims: Identify transcriptomic signatures and trajectories of VSMCs induced by the 9p21.3 risk haplotype Methods: We have used iPSCs from individuals carrying the risk and non-risk haplotype at 9p21.3 and isogenic knockout lines. After VSMCs differentiation induction, mature VSMCs were used for single cell RNA sequencing using 10X Chromium platform. Results: Our analysis revealed that the 9p21.3 risk haplotype prompts VSMCs to acquire a novel cellular state showing reduced plasticity and divergent from other VSMC states previously linked to CAD. We found markers of alternative lineages, including osteogenic and neuronal, coupled with altered expression of genes within other CAD loci. We identified a set of new molecular markers crucial to define risk-associated VSMCs and uncovered their upstream regulators. Conclusions: Our study provides insights into CAD pathogenesis driven by the 9p21.3 risk locus and identified new gene regulatory networks essential for maintaining the normal functionality of the muscle layer of the arteries. Leveraging the power of iPSCs we present novel concepts about the biology of VSMCs and shed light on the impact of CAD genetic risk factors in preserving a healthy cellular state within the vasculature.

  PublisherDOI

• Heterogeneous expression of alternatively spliced lncRNA mediates vascular smooth cell plasticity

  Proceedings of the National Academy of Sciences · 2023 · 11 citations

  • Biology
  • Cell biology
  • Genetics

  suppressed adhesion, contractility, and αSMA expression. These data suggest that variable lncRNA penetrance may drive mixed functional outcomes that confound pathology.

  PublisherOA PDFDOI

• High shear stress enhances endothelial permeability in the presence of the risk haplotype at 9p21.3

  APL Bioengineering · 2021 · 6 citations

  • Biology
  • Immunology
  • Cell biology

  Single nucleotide polymorphisms (SNPs) are exceedingly common in non-coding loci, and while they are significantly associated with a myriad of diseases, their specific impact on cellular dysfunction remains unclear. Here, we show that when exposed to external stressors, the presence of risk SNPs in the 9p21.3 coronary artery disease (CAD) risk locus increases endothelial monolayer and microvessel dysfunction. Endothelial cells (ECs) derived from induced pluripotent stem cells of patients carrying the risk haplotype (R/R WT) differentiated similarly to their non-risk and isogenic knockout (R/R KO) counterparts. Monolayers exhibited greater permeability and reactive oxygen species signaling when the risk haplotype was present. Addition of the inflammatory cytokine TNFα further enhanced EC monolayer permeability but independent of risk haplotype; TNFα also did not substantially alter haplotype transcriptomes. Conversely, when wall shear stress was applied to ECs in a microfluidic vessel, R/R WT vessels were more permeable at lower shear stresses than R/R KO vessels. Transcriptomes of sheared cells clustered more by risk haplotype than by patient or clone, resulting in significant differential regulation of EC adhesion and extracellular matrix genes vs static conditions. A subset of previously identified CAD risk genes invert expression patterns in the presence of high shear concomitant with altered cell adhesion genes, vessel permeability, and endothelial erosion in the presence of the risk haplotype, suggesting that shear stress could be a regulator of non-coding loci with a key impact on CAD.

  PublisherOA PDFDOI

• Metabolic Dysregulation of the Lysophospholipid/Autotaxin Axis in the Chromosome 9p21 Gene SNP rs10757274

  Circulation Genomic and Precision Medicine · 2020 · 12 citations

  • Biology
  • Genetics

  BACKGROUND: Common chromosome 9p21 single nucleotide polymorphisms (SNPs) increase coronary heart disease risk, independent of traditional lipid risk factors. However, lipids comprise large numbers of structurally related molecules not measured in traditional risk measurements, and many have inflammatory bioactivities. Here, we applied lipidomic and genomic approaches to 3 model systems to characterize lipid metabolic changes in common Chr9p21 SNPs, which confer ≈30% elevated coronary heart disease risk associated with altered expression of ANRIL, a long ncRNA. METHODS: Untargeted and targeted lipidomics was applied to plasma from NPHSII (Northwick Park Heart Study II) homozygotes for AA or GG in rs10757274, followed by correlation and network analysis. To identify candidate genes, transcriptomic data from shRNA downregulation of ANRIL in HEK-293 cells was mined. Transcriptional data from vascular smooth muscle cells differentiated from induced pluripotent stem cells of individuals with/without Chr9p21 risk, nonrisk alleles, and corresponding knockout isogenic lines were next examined. Last, an in-silico analysis of miRNAs was conducted to identify how ANRIL might control lysoPL (lysophosphospholipid)/lysoPA (lysophosphatidic acid) genes. RESULTS: The in-silico analysis identified 4 ANRIL-regulated miRNAs that control lysoPL genes as miR-186-3p, miR-34a-3p, miR-122-5p, and miR-34a-5p. CONCLUSIONS: A Chr9p21 risk SNP associates with complex alterations in immune-bioactive phospholipids and their metabolism. Lipid metabolites and genomic pathways associated with coronary heart disease pathogenesis in Chr9p21 and ANRIL-associated disease are demonstrated.

  PublisherOA PDFDOI

• Mechanical activation of noncoding-RNA-mediated regulation of disease-associated phenotypes in human cardiomyocytes

  Nature Biomedical Engineering · 2019-01-28 · 41 citations

  articleOpen access
  PublisherOA PDFDOI

• Unveiling the Role of the Most Impactful Cardiovascular Risk Locus through Haplotype Editing

  Cell · 2018-12-01 · 129 citations

  articleOpen access1st authorCorresponding
  PublisherOA PDFDOI

• Abstract 289: Improved Disease Modeling Reveals 9p21 Risk Allele Regulates Connexins to Induce Arrhythmic Phenotypes

  Circulation Research · 2017-07-21

  article

  Genome-wide association studies have identified single nucleotide polymorphisms (SNPs) in the non-coding 9p21 gene locus associated with increased risk of coronary artery disease (CAD) and myocardial infarction (MI). In addition, SNP correlations with sudden and arrhythmic death, even after accounting for patient and family history for CAD and MI, suggest an altered cardiac remodeling response. However, little is known about a possible cardiac phenotype as studies have largely focused on its effect on CAD and have trouble describing regulation with non-coding loci. Using induced pluripotent stem cell-derived CMs from patients that are homozygous risk/risk (R/R) and non-risk/non-risk (N/N) for 9p21 SNPs, we assessed cardiomyocyte (CM) function when cultured on hydrogels capable of mimicking the fibrotic stiffening associated with disease post-heart attack, i.e. stiffening from 10 kiloPascals (kPa) to 50 kPa. While all CMs independent of genotype beat synchronously on soft matrices, R/R CMs cultured on dynamically stiffened hydrogels exhibited asynchronous contractions versus N/N CMs in the same conditions. Dynamic stiffening reduced connexin 43 expression and gap junction assembly in R/R CMs but not N/N CMs. To eliminate patient-to-patient variability, we created an isogenic line by deleting the 9p21 locus from a R/R patient, i.e. R/R KO. R/R KO CMs maintained synchronous contractions and organized connexin 43 junctions after stiffening. The 9p21 locus suppresses the activity of the cell cycle regulator CDKN2A. p16, a protein produced by CDKN2A, prevents JNK phosphorylation (p-JNK), which in turn reduces gap junction expression in CMs and contributes to the development of arrhythmias in rabbit myocardium in response to stress. We observed that treatment with the p-JNK antagonist SP600125 after stiffening restored synchronous contractions and organized gap junction assembly to R/R CM. As a non-coding locus, 9p21 appears to repress connexin transcription, but only when the niche is stiffened as in disease. These data are the first to demonstrate that disease-specific niche remodeling can differentially affect CM function depending on SNPs within a non-coding locus.

  PublisherDOI

Frequent coauthors

• Adam J. Engler

  University of California, San Diego

  11 shared

• Kristin K. Baldwin

  Columbia University Irving Medical Center

  8 shared

• William C. Ferguson

  7 shared

• Aditya Kumar

  La Jolla Bioengineering Institute

  6 shared

• Eric J. Topol

  Scripps Clinic

  6 shared

• Ali Torkamani

  Scripps Health

  4 shared

• Elsa Salido

  University of Wisconsin–Madison

  3 shared

• Kevin Tenerelli

  University of California, San Diego

  3 shared

Labs

• Lo Sardo laboratoryPI

Education

• PhD in Pharmaceutical Biotechnology, Department of Pharmacological Sciences

  University of Milan

  2010

• MSc in Pharmaceutical Biotechnology, Department of Pharmacological Sciences

  University of Milan

  2006