Biology subjects

Hamdan, S.

Publications and source records attributed to Hamdan, S..

4 recordsLinked to original sources

bioRxiv2022

Modulation of the microhomology-mediated end joining pathway suppresses large deletions and enhances homology-directed repair following CRISPR-Cas9-induced DNA breaks

CRISPR-Cas9, an efficient genome editing tool, has been widely used in research and holds great promise in the clinic. However, large unintended rearrangements of the genome occur frequently after CRISPR-Cas9 editing and their potential risk cannot be ignored. In this study, we detected large deletions (LDs) induced by CRISPR-Cas9 in human embryonic stem cells (hESCs) and found the microhomology end joining (MMEJ) DNA repair pathway plays a predominant role in LD. We genetically targeted PARP1, RPA, POLQ and LIG3, which play critical roles in MMEJ, during CRISPR-Cas9 editing. By analyzing LD events in two independent gene loci, CD9 and PIGA, using flow cytometry and long-read individual molecule sequencing (IDMseq), we showed that knocking down PARP1 and LIG3 does not alter the frequency of Cas9-induced LD, while knocking down or inhibiting POLQ dramatically reduces LD. Knocking down RPA increases LD frequency, and overexpression of RPAs consistently reduces LD frequency. Interestingly, small-molecule inhibition of POLQ and delivery of recombinant RPA proteins also dramatically increase the efficiency of homology-directed repair (HDR). In conclusion, RPA and POLQ play opposite roles in Cas9-induced LD, modulation of POLQ and RPA can reduce LD and improve HDR, thus holding promise for safe and precise genome editing.

molecular biology↗

bioRxiv2022

Accurate sex prediction of cisgender and transgender individuals without brain size bias

Brain size differs substantially between human males and females. This difference in total intracranial volume (TIV) can cause bias when employing machine-learning approaches for the investigation of sex differences in brain morphology. TIV-biased models will likely not capture actual qualitative sex differences in brain organization but rather learn to classify an individuals sex based on brain size differences, thus leading to spurious and misleading conclusions, for example when comparing brain morphology between cisgender- and transgender individuals. Here, TIV bias in sex classification models applied to cis- and transgender individuals was systematically investigated by controlling for brain size either through featurewise confound removal or by matching training samples for TIV. Our results provide evidence that non-TIV-biased models can classify the sex of both cis- and transgender individuals with high accuracy, highlighting the importance of appropriate modelling to avoid bias in automated decision making. TeaserAccurate non-biased structural sex classification in cis- and transgender individuals by matching training samples for TIV

neuroscience↗

bioRxiv2022

Urine-derived exosomes from individuals with IPF carry pro-fibrotic cargo.

BackgroundMicroRNAs (miRNA) and other components contained in extracellular vesicles may reflect the presence of a disease. Lung tissue, sputum and sera of individuals with idiopathic pulmonary fibrosis (IPF) show alterations in miRNA expression. We designed this study to test whether urine and/or tissue derived exosomal miRNAs from individuals with IPF carry cargo that can promote fibrosis. MethodsExosomes were isolated from urine (U-IPFexo), lung tissue myofibroblasts (MF-IPFexo), serum from individuals with IPF (n=16) and age/sex-matched controls without lung disease (n=10). We analyzed microRNA expression of isolated exosomes and their in vivo bio-distribution. We investigated the effect on ex vivo skin wound healing and in in vivo mouse lung models. ResultsU-IPFexo or MF-IPFexo expressed miR let-7d, miR-29a-5p, miR 181b-3p and miR-199a-3p consistent with previous reports of miRNA expression obtained from lung tissue/sera from patients with IPF. In vivo bio-distribution experiments detected bioluminescent exosomes in the lung of normal C57Bl6 mice within 5 minutes after intravenous infusion, followed by distribution to other organs irrespective of exosome source. Exosomes labeled with gold nanoparticles and imaged by transmission electron microscopy were visualized in alveolar epithelial type I and type II cells. Treatment of human and mouse lung punches obtained from control, non-fibrotic lungs with either U-IPFexo or MF-IPFexo produced a fibrotic phenotype. A fibrotic phenotype was also induced in a human ex vivo skin model and in in vivo lung models. ConclusionsOur results provide evidence of a systemic feature of IPF whereby exosomes contain pro-fibrotic miRNAs when obtained from a fibrotic source and interfere with response to tissue injury as measured in skin and lung models. FundingThis work was supported in part by Lester and Sue Smith Foundation and The Samrick Family Foundation and NIH grants R21 AG060338 (SE and MKG), U01 DK119085 (IP, RS, MTC).

cell biology↗

bioRxiv2021

Aspirin Protective Effect Against Cyclophosphamide Hematological Toxicity In Experimental Animals

Bone marrow toxicity is the most important factor limiting the use of cytotoxic drugs like alkylating agents in cancer treatment. Recently PG synthase enzyme inhibitors have been reported to potentiate the cytotoxic effects of these agents on cancer cells but little is known if they can affect the toxicity of these agents on bone marrow or other tissues. Cyclophosphamide is one of the most commonly used alkylating agent. In the present work, the effect of these PG synthase enzyme inhibitors, aspirin on cyclophosphamide myelotoxicity was determined employing the peripheral blood count to reflect bone marrow injury. The effect on body weight changes caused by cyclophosphamide was also determined. O_LICyclophosphamide in doses of 25, 50 and 75 mg/kg i. v. produced as a dose dependent reduction in total WBC count, granulocyte, non granulocyte, and Hb% which was maximum on second day after injection and still present on 5th day post injection. It also produced a dose dependent reduction in body weight on day 5 after injection. C_LIO_LIAspirin in doges of 75, 150 and 300 mg/kg i. m. protected against the reduction in WBC counts measured for 5 days after injection of cyclophosphamide (50 mg/kg). This protection was not dose dependent, though it was more optimum with 300 mg/kg and disappeared largely when a dose of 450 mg/kg was used. Aspirin did not prevent the changes in Hb% but retard the reduction in body weight caused by cyclophosphamide. C_LIO_LIIt is concluded that aspirin can help to reduce injury and enhance recovery from bone marrow toxicity caused by cytotoxic agents such as the alkylating drugs cyclophosphamide for which no specific antidote is available. Aspirin produces this effect possibly by eliminating the harmful inhibitory effect of excess PGs or leukotrienes, released by bone marrow injury on growth factors of haemopoietic progenitor cells. C_LI The magnitude of this protection on WBC counts does not seem to differ between either PG synthase enzyme inhibitors or steroids when used alone or in combination although a synergistic effect in protecting erythropoiesis is observed.

pharmacology and toxicology↗