Use of mouse hematopoietic stem and progenitor cells to treat acute kidney injury

New and effective treatment for acute kidney injury remains a challenge. Here, we induced mouse hematopoietic stem and progenitor cells (HSPC) to differentiate into cells that partially resemble a renal cell phenotype and tested their therapeutic potential. We sequentially treated HSPC with a combination of protein factors for 1 wk to generate a large number of cells that expressed renal developmentally regulated genes and protein. Cell fate conversion was associated with increased histone acetylation on promoters of renal-related genes. Further treatment of the cells with a histone deacetylase inhibitor improved the efficiency of cell conversion by sixfold. Treated cells formed tubular structures in three-dimensional cultures and were integrated into tubules of embryonic kidney organ cultures. When injected under the renal capsule, they integrated into renal tubules of postischemic kidneys and expressed the epithelial marker E-cadherin. No teratoma formation was detected 2 and 6 mo after cell injection, supporting the safety of using these cells. Furthermore, intravenous injection of the cells into mice with renal ischemic injury improved kidney function and morphology by increasing endogenous renal repair and decreasing tubular cell death. The cells produced biologically effective concentrations of renotrophic factors including VEGF, IGF-1, and HGF to stimulate epithelial proliferation and tubular repair. Our study indicates that hematopoietic stem and progenitor cells can be converted to a large number of renal-like cells within a short period for potential treatment of acute kidney injury.

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Faculty Opinions recommendation of Use of mouse hematopoietic stem and progenitor cells to treat acute kidney injury.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.13296966.14657068 ◽

2011 ◽

Author(s):

Angela Wandinger-Ness ◽

Heather Ward

Keyword(s):

Acute Kidney Injury ◽

Progenitor Cells ◽

Kidney Injury ◽

Hematopoietic Stem ◽

Stem And Progenitor Cells

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S. Aureus Cas9 Leads to Higher Sickle Mutation Correction Compared to S. Pyogenes Cas9 in Patient Hematopoietic Stem and Progenitor Cells

Blood ◽

10.1182/blood-2021-150830 ◽

2021 ◽

Vol 138 (Supplement 1) ◽

pp. 1861-1861

Author(s):

Byoungyong Yoo ◽

So Hyun Julie Park Park ◽

Yankai Zhang ◽

Vivien A. Sheehan ◽

Gang Bao

Keyword(s):

Progenitor Cells ◽

Research Funding ◽

Gene Editing ◽

Cell Phenotype ◽

High Morbidity ◽

Gene Correction ◽

Hematopoietic Stem ◽

Nucleotide Mutation ◽

Stem And Progenitor Cells ◽

Donor Template

Abstract Introduction: Sickle cell disease (SCD) is a red blood cell disorder caused by a single nucleotide mutation in the β-globin gene (HBB). Allogeneic hematopoietic stem cell transplantation (HSCT) is the only available cure, but is available to only a minority of patients and can be associated with high morbidity and mortality. CRISPR/Cas9 mediated genome editing may provide a permanent cure for SCD patients by correcting the sickle mutation in HBB in hematopoietic stem and progenitor cells (HSPCs). Previously, we achieved ~39% sickle mutation correction in SCD HSPCs by delivering S. pyogenes (Spy) Cas9/R-66S gRNA as ribonucleoprotein (RNP) and single-stranded oligodeoxynucleotides (ssODN) corrective donor template. S. aureus (Sau) Cas9 has potentially advantageous properties to improve therapeutic gene editing efficiency and safety, including smaller size allowing for efficient in vivo delivery and longer Protospacer Adjacent Motif (PAM) sequence for higher specificity. However, although in general, the cutting efficiency of SauCas9 is lower than SpyCas9, the differences in gene correction and other gene-editing outcomes between SpyCas9 and SauCas9 have not been well studied. Methods: With our R-66S gRNA sequence targeting the sickle mutation, the PAM sequence of SauCas9 (NGGRRT) is mutually permissive with that of SpyCas9 (NGG), allowing the same sequence to be targeted by both Cas9 nucleases. We delivered R-66S gRNA with SpyCas9 and SauCas9 respectively as RNP, along with corrective ssODN donor template into SCD HSPCs. We analyzed sickle mutation correction rate and small insertions and deletions (INDELs) profile by Next Generation Sequencing (NGS). Results/discussions: We found that although the INDEL rate of SpyCas9 is higher than SauCas9 at the same molar concentration of RNP, SauCas9 gave 43% sickle mutation correction, slightly higher than SpyCas9 (39%), demonstrating efficient homology-directed repair (HDR) mediated gene correction by SauCas9. To further investigate the potential for clinical translation, we will perform in-depth efficiency and safety characterization comparing SauCas9 and SpyCas9 mediated sickle mutation correction therapy in SCD HSPCs. Conclusion: In this work, we showed that, compared with the highly-optimized and widely-used SpyCas9, SauCas9 leads to a higher sickle mutation correction in SCD HSPCs, demonstrating the therapeutic potential of SauCas9 for treating SCD. We will further investigate the efficiency and safety of gene-edited therapy mediated by these two Cas9 orthologs, including in-depth characterization of off-target effects, chromosomal rearrangement and aberrations, and large genomic modifications. We will differentiate gene-corrected SCD HSPCs to study erythropoiesis and red cell phenotype, including normal hemoglobin production and reduced sickling under hypoxic conditions. Lastly, we will evaluate the engraftment efficiency of gene-edited cells in Nonirradiated NOD,B6.SCID Il2rγ -/- Kit (W41/W41) (NBSGW) mice that support the engraftment of human hematopoietic stem cells. Disclosures Sheehan: Forma Therapeutics: Research Funding; Beam Therapeutics: Research Funding; Novartis: Research Funding.

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Skeletal Muscle-Derived Stem/Progenitor Cells: A Potential Strategy for the Treatment of Acute Kidney Injury

Stem Cells International ◽

10.1155/2016/9618480 ◽

2016 ◽

Vol 2016 ◽

pp. 1-13 ◽

Cited By ~ 7

Author(s):

Egle Pavyde ◽

Romaldas Maciulaitis ◽

Mykolas Mauricas ◽

Gintaras Sudzius ◽

Ernesta Ivanauskaite Didziokiene ◽

...

Keyword(s):

Skeletal Muscle ◽

Acute Kidney Injury ◽

Progenitor Cells ◽

Therapeutic Potential ◽

Kidney Injury ◽

Population Doubling ◽

Population Doubling Time ◽

Isolation Technique ◽

Time Flow ◽

Potential Strategy

Skeletal muscle-derived stem/progenitor cells (MDSPCs) have been thoroughly investigated and already used in preclinical studies. However, therapeutic potential of MDSPCs isolated using preplate isolation technique for acute kidney injury (AKI) has not been evaluated. We aimed to characterize rat MDSPCs, compare them with bone marrow mesenchymal stem cells (BM-MSCs), and evaluate the feasibility of MDSPCs therapy for gentamicin-induced AKI in rats. We have isolated and characterized rat MDSPCs and BM-MSCs. Characteristics of rat BM-MSCs and MDSPCs were assessed by population doubling time, flow cytometry, immunofluorescence staining, RT-PCR, and multipotent differentiation capacity. Gentamicin-induced AKI model in rat was used to examine MDSPCs therapeutic effect. Physiological and histological kidney parameters were determined. MDSPCs exhibited similar immunophenotype, stem cell gene expression, and multilineage differentiation capacities as BM-MSCs, but they demonstrated higher proliferation rate. Single intravenous MDSPCs injection accelerated functional and morphological kidney recovery, as reflected by significantly lower serum creatinine levels, renal injury score, higher urinary creatinine, and GFR levels. PKH-26-labeled MDSPCs were identified within renal cortex 1 and 2 weeks after cell administration, indicating MDSPCs capacity to migrate and populate renal tissue. In conclusion, MDSPCs are capable of mediating functional and histological kidney recovery and can be considered as potential strategy for AKI treatment.

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Inhibition of HDAC6 protects against rhabdomyolysis-induced acute kidney injury

AJP Renal Physiology ◽

10.1152/ajprenal.00546.2016 ◽

2017 ◽

Vol 312 (3) ◽

pp. F502-F515 ◽

Cited By ~ 31

Author(s):

Yingfeng Shi ◽

Liuqing Xu ◽

Jinhua Tang ◽

Lu Fang ◽

Shuchen Ma ◽

...

Keyword(s):

Acute Kidney Injury ◽

Therapeutic Potential ◽

Apoptotic Cell ◽

Kidney Injury ◽

Acute Injury ◽

Tubular Damage ◽

Renal Tubules ◽

Tubular Cells ◽

Renal Tubular Cells ◽

Renal Tubular

Histone deacetylase 6 (HDAC6) inhibition has been reported to protect against ischemic stroke and prolong survival after sepsis in animal models. However, it remains unknown whether HDAC6 inhibition offers a renoprotective effect after acute kidney injury (AKI). In this study, we examined the effect of tubastatin A (TA), a highly selective inhibitor of HDAC6, on AKI in a murine model of glycerol (GL) injection-induced rhabdomyolysis. Following GL injection, the mice developed severe acute tubular injury as indicated by renal dysfunction; expression of neutrophil gelatinase-associated lipocalin (NGAL), an injury marker of renal tubules; and an increase of TdT-mediated dUTP nick-end labeling (TUNEL)-positive tubular cells. These changes were companied by increased HDAC6 expression in the cytoplasm of renal tubular cells. Administration of TA significantly reduced serum creatinine and blood urea nitrogen levels as well as attenuated renal tubular damage in injured kidneys. HDAC6 inhibition also resulted in decreased expression of NGAL, reduced apoptotic cell, and inactivated caspase-3 in the kidney after acute injury. Moreover, injury to the kidney increased phosphorylation of nuclear factor (NF)-κB and expression of multiple cytokines/chemokines including tumor necrotic factor-α and interleukin-6 and monocyte chemoattractant protein-1, as well as macrophage infiltration. Treatment with TA attenuated all those responses. Finally, HDAC6 inhibition reduced the level of oxidative stress by suppressing malondialdehyde (MDA) and preserving expression of superoxide dismutase (SOD) in the injured kidney. Collectively, these data indicate that HDAC6 contributes to the pathogenesis of rhabdomyolysis-induced AKI and suggest that HDAC6 inhibitors have therapeutic potential for AKI treatment.

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Mitophagy in Acute Kidney Injury and Kidney Repair

Cells ◽

10.3390/cells9020338 ◽

2020 ◽

Vol 9 (2) ◽

pp. 338 ◽

Cited By ~ 14

Author(s):

Ying Wang ◽

Juan Cai ◽

Chengyuan Tang ◽

Zheng Dong

Keyword(s):

Acute Kidney Injury ◽

Kidney Disease ◽

Molecular Mechanisms ◽

Therapeutic Potential ◽

Kidney Injury ◽

Future Research ◽

Rapid Decline ◽

Renal Tubules ◽

Renal Tubular Cells

Acute kidney injury (AKI) is a major kidney disease characterized by rapid decline of renal function. Besides its acute consequence of high mortality, AKI has recently been recognized as an independent risk factor for chronic kidney disease (CKD). Maladaptive or incomplete repair of renal tubules after severe or episodic AKI leads to renal fibrosis and, eventually, CKD. Recent studies highlight a key role of mitochondrial pathology in AKI development and abnormal kidney repair after AKI. As such, timely elimination of damaged mitochondria in renal tubular cells represents an important quality control mechanism for cell homeostasis and survival during kidney injury and repair. Mitophagy is a selective form of autophagy that selectively removes redundant or damaged mitochondria. Here, we summarize our recent understanding on the molecular mechanisms of mitophagy, discuss the role of mitophagy in AKI development and kidney repair after AKI, and present future research directions and therapeutic potential.

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Combined single-cell gene and isoform expression analysis in haematopoietic stem and progenitor cells

10.1101/2020.04.06.027474 ◽

2020 ◽

Cited By ~ 1

Author(s):

Laura Mincarelli ◽

Vladimir Uzun ◽

Stuart A. Rushworth ◽

Wilfried Haerty ◽

Iain C. Macaulay

Keyword(s):

Single Cell ◽

Progenitor Cells ◽

Cell Types ◽

Cell Phenotype ◽

Hematopoietic Stem ◽

Long Read ◽

Stem And Progenitor Cells ◽

Alternatively Spliced ◽

Isoform Expression

AbstractSingle-cell RNA sequencing (scRNA-seq) enables gene expression profiling and characterization of novel cell types within heterogeneous cell populations. However, most approaches cannot detect alternatively spliced transcripts, which can profoundly shape cell phenotype by generating functionally distinct proteins from the same gene. Here, we integrate short- and long-read scRNA-seq of hematopoietic stem and progenitor cells to characterize changes in cell type abundance, gene and isoform expression during differentiation and ageing.

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