Utilization of nicking properties of CRISPR-Cas12a effector for genome editing.

The CRISPR-Cas nickase system for genome editing has attracted considerable attention owing to its safety, efficiency, and versatility. Although alternative effectors to Cas9 have the potential to expand the scope of genome editing, their application has not been optimized. Herein, we used an enhanced CRISPR-Cas12a nickase system to induce mutations by targeting genes in a human-derived cell line. The optimized CRISPR-Cas12a nickase system effectively introduced mutations into target genes under a specific directionality and distance between nickases. In particular, the single-mode Cas12a nickase system can induce the target-specific mutations with less DNA double-strand breaks. By inducing mutations in the Thymine-rich target genes in single- or dual-mode, Cas12a nickase compensates the limitations of Cas9 nickase and is expected to contribute to the development of future genome editing technologies.

Identification of microRNAs Derived from Transposable Elements in the Macaca mulatta (Rhesus Monkey) Genome.

Transposable elements (TEs) are mobile DNA entities that can move within the host genome. Over long periods of evolutionary time, TEs are typically silenced via the accumulation of mutations in the genome, ultimately resulting in their immobilization. However, they still play an important role in the host genome by acting as regulatory elements. They influence host transcription in various ways, one of which as the origin of the generation of microRNAs (miRNAs), which are so-called miRNAs derived from TEs (MDTEs). miRNAs are small non-coding RNAs that are involved in many biological processes by regulating gene expression at the post-transcriptional level. Here, we identified MDTEs in the Macaca mulatta (rhesus monkey) genome, which is phylogenetically close species to humans, based on the genome coordinates of miRNAs and TEs. The expression of 5 out of 17 MDTEs that were exclusively registered in M. mulatta from the miRBase database (v22) was examined via quantitative polymerase chain reaction (qPCR). Moreover, Gene Ontology analysis was performed to examine the functional implications of the putative target genes of the five MDTEs.

Analysis of Factors Affecting Postoperative Opioid Requirement in Pediatric Patients Undergoing Pectus Excavatum Repair with Multimodal Analgesic Management.

Children with pectus excavatum are treated with surgical repair in a procedure known as minimally invasive repair of pectus excavatum (MIRPE). MIRPE causes considerable postoperative pain, resulting in the administration of a substantial dose of opioids. This study aimed to identify perioperative factors that influence the requirement for opioids in children undergoing MIRPE. Retrospective data from children who underwent MIRPE were analyzed. A multimodal analgesic protocol was implemented with a continuous wound infiltration system and administration of non-opioid analgesics. Intravenous opioid analgesics were administered if the pain score was greater than 4. The cumulative opioid use was assessed by calculating the morphine equivalent dose at 6, 24, and 48 h after surgery. Perioperative factors affecting the postoperative opioid use were identified with multiple linear regression analyses. This study included 527 children aged 3-6 years, with a mean age of 3.9 years. Symmetrically depressed chest walls, a lower Haller index, and a lower revised depression index were found to be associated with decreased postoperative opioids. Boys required higher opioid doses than girls. Longer pectus bars (10 inches versus 9 inches) were associated with increased opioid use. Severity indices, gender, and the length of pectus bars influence postoperative opioid requirement in children undergoing MIRPE surgery with multimodal analgesia.

Rho MultiBinder, a fluorescent biosensor that reports the activity of multiple GTPases.

Imaging two or more fluorescent biosensors in the same living cell can reveal the spatiotemporal coordination of protein activities. However, using multiple Förster resonance energy transfer (FRET) biosensors together is challenging due to toxicity and the need for orthogonal fluorophores. Here we generate a biosensor component that binds selectively to the activated conformation of three different proteins. This enabled multiplexed FRET with fewer fluorophores, and reduced toxicity. We generated this MultiBinder (MB) reagent for the GTPases RhoA, Rac1, and Cdc42 by combining portions of the downstream effector proteins Pak1 and Rhotekin. Using FRET between mCherry on the MB and YPet or mAmetrine on two target proteins, the activities of any pair of GTPases could be distinguished. The MB was used to image Rac1 and RhoA together with a third, dye-based biosensor for Cdc42. Quantifying effects of biosensor combinations on the frequency, duration, and velocity of cell protrusions and retractions demonstrated reduced toxicity. Multiplexed imaging revealed signaling hierarchies between the three proteins at the cell edge where they regulate motility.

Expression of the melatonergic system during meiotic maturation of cynomolgus monkey cumulus-oocyte complexes.

BACKGROUND: Melatonin is a multifunctional hormone synthesized in the pineal gland and peripheral reproductive tissues that regulates many biological processes. There is increasing evidence for a role of melatonin in oocyte maturation and embryonic development in various mammals. However, no study has reported evidence for the existence of melatonergic system, such as melatonin synthesis enzymes, melatonin membrane receptors, or melatonin binding sites in non-human primate cumulus-oocyte complexes (COCs). METHODS: Reverse transcription polymerase chain reaction (RT-PCR) and immunocytochemistry were performed to detect transcripts and proteins of the rate-limiting enzyme in melatonin synthesis (arylalkylamine N-acetyltransferase, AANAT), melatonin membrane receptors (MT1 and MT2), and a melatonin binding site (NRH: quinone oxidoreductase 2, NQO2) in cynomolgus monkey COCs. RESULTS: RT-PCR analyses revealed the presence of AANAT, MT1, MT2, and NQO2 transcripts in granulosa cells, germinal vesicle (GV)- and metaphase II (MII)-stage cumulus cells, and oocytes. Immunocytochemistry revealed the presence of AANAT, MT1, MT2, and NQO2 proteins in GV- and MII-stage COCs. CONCLUSIONS: Our results provide the first evidence for the existence of the rate-limiting enzyme required for melatonin synthesis, melatonin membrane receptors, and a melatonin binding site in non-human primate COCs.

Fermented Soybean Paste Attenuates Biogenic Amine-Induced Liver Damage in Obese Mice.

Biogenic amines are cellular components produced by the decarboxylation of amino acids; however, excessive biogenic amine production causes adverse health problems. The relationship between hepatic damage and biogenic amine levels in nonalcoholic fatty liver disease (NAFLD) remains unclear. In this study, mice were fed a high-fat diet (HFD) for 10 weeks to induce obesity, presenting early-stage of NAFLD. We administered histamine (20 mg/kg) + tyramine (100 mg/kg) via oral gavage for 6 days to mice with HFD-induced early-stage NAFLD. The results showed that combined histamine and tyramine administration increased cleaved PARP-1 and IL-1ß in the liver, as well as MAO-A, total MAO, CRP, and AST/ALT levels. In contrast, the survival rate decreased in HFD-induced NAFLD mice. Treatment with manufactured or traditional fermented soybean paste decreased biogenically elevated hepatic cleaved PARP-1 and IL-1ß expression and blood plasma MAO-A, CRP, and AST/ALT levels in HFD-induced NAFLD mice. Additionally, the biogenic amine-induced reduction in survival rate was alleviated by fermented soybean paste in HFD-induced NAFLD mice. These results show that biogenic amine-induced liver damage can be exacerbated by obesity and may adversely affect life conservation. However, fermented soybean paste can reduce biogenic amine-induced liver damage in NAFLD mice. These results suggest a beneficial effect of fermented soybean paste on biogenic amine-induced liver damage and provide a new research perspective on the relationship between biogenic amines and obesity.

Determination of the Unilaterally Damaged Region May Depend on the Asymmetry of Carotid Blood Flow Velocity in Hemiparkinsonian Monkey: A Pilot Study.

The hemiparkinsonian nonhuman primate model induced by unilateral injection of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) into the carotid artery is used to study Parkinson's disease. However, there have been no studies that the contralateral distribution of MPTP via the cerebral collateral circulation is provided by both the circle of Willis (CoW) and connections of the carotid artery. To investigate whether MPTP-induced unilaterally damaged regions were determined by asymmetrical cerebral blood flow, the differential asymmetric damage of striatal subregions, and examined structural asymmetries in a circle of Willis, and blood flow velocity of the common carotid artery were observed in three monkeys that were infused with MPTP through the left internal carotid artery. Lower flow velocity in the ipsilateral common carotid artery and a higher ratio of ipsilateral middle cerebral artery diameter to anterior cerebral artery diameter resulted in unilateral damage. Additionally, the unilateral damaged monkey observed the apomorphine-induced contralateral rotation behavior and the temporary increase of plasma RANTES. Contrastively, higher flow velocity in the ipsilateral common carotid artery was observed in the bilateral damaged monkey. It is suggested that asymmetry of blood flow velocity and structural asymmetry of the circle of Willis should be taken into consideration when establishing more efficient hemiparkinsonian nonhuman primate models.

Highly specific chimeric DNA-RNA-guided genome editing with enhanced CRISPR-Cas12a system.

The clustered regularly interspaced short palindromic repeats (CRISPR)-Cas12a system is composed of a Cas12a effector that acts as a DNA-cleaving endonuclease and a crispr RNA (crRNA) that guides the effector to the target DNA. It is considered a key molecule for inducing target-specific gene editing in various living systems. Here, we improved the efficiency and specificity of the CRISPR-Cas12a system through protein and crRNA engineering. In particular, to optimize the CRISPR-Cas12a system at the molecular level, we used a chimeric DNA-RNA guide chemically similar to crRNA to maximize target sequence specificity. Compared with the wild-type (wt)-Cas12a system, when using enhanced Cas12a system (en-Cas12a), the efficiency and target specificity improved on average by 2.58 and 2.77 times, respectively. In our study, when the chimeric DNA-RNA-guided en-Cas12a effector was used, the gene-editing efficiency and accuracy were simultaneously increased. These findings could contribute to highly accurate genome editing, such as human gene therapy, in the near future.

Hsa-miR-422a Originated from Short Interspersed Nuclear Element Increases ARID5B Expression by Collaborating with NF-E2.

MicroRNAs (miRNAs) are a class of small non-coding RNAs that regulate the expression of target messenger RNA (mRNA) complementary to the 3' untranslated region (UTR) at the post-transcriptional level. Hsa-miR-422a, which is commonly known as miRNA derived from transposable element (MDTE), was derived from short interspersed nuclear element (SINE). Through expression analysis, hsa-miR-422a was found to be highly expressed in both the small intestine and liver of crab-eating monkey. AT-Rich Interaction Domain 5 B (ARID5B) was selected as the target gene of hsa-miR-422a, which has two binding sites in both the exon and 3'UTR of ARID5B. To identify the interaction between hsa-miR-422a and ARID5B, a dual luciferase assay was conducted in HepG2 cell line. The luciferase activity of cells treated with the hsa-miR-422a mimic was upregulated and inversely downregulated when both the hsa-miR-422a mimic and inhibitor were administered. Nuclear factor erythroid-2 (NF-E2) was selected as the core transcription factor (TF) via feed forward loop analysis. The luciferase expression was downregulated when both the hsa-miR-422a mimic and siRNA of NF-E2 were treated, compared to the treatment of the hsa-miR-422a mimic alone. The present study suggests that hsa-miR-422a derived from SINE could bind to the exon region as well as the 3'UTR of ARID5B. Additionally, hsa-miR-422a was found to share binding sites in ARID5Bwith several TFs, including NF-E2. The hsa-miR-422a might thus interact with TF to regulate the expression of ARID5B, as demonstrated experimentally. Altogether, hsa-miR-422a acts as a super enhancer miRNA of ARID5Bby collaborating with TF and NF-E2.

Expansion of the prime editing modality with Cas9 from Francisella novicida.

Prime editing can induce a desired base substitution, insertion, or deletion in a target gene using reverse transcriptase after nick formation by CRISPR nickase. In this study, we develop a technology that can be used to insert or replace external bases in the target DNA sequence by linking reverse transcriptase to the Francisella novicida Cas9, which is a CRISPR-Cas9 ortholog. Using FnCas9(H969A) nickase, the targeting limitation of existing Streptococcus pyogenes Cas9 nickase [SpCas9(H840A)]-based prime editing is dramatically extended, and accurate prime editing is induced specifically for the target genes in human cell lines.

Intraoperative Glycemic Variability and Mean Glucose are Predictors for Postoperative Delirium After Cardiac Surgery: A Retrospective Cohort Study.

PURPOSE: Postoperative delirium (POD) is a common but serious complication after cardiac surgery and is associated with various short- and long-term outcomes. In this study, we investigated the effects of intraoperative glycemic variability (GV) and other glycemic variables on POD after cardiac surgery. PATIENTS AND METHODS: A retrospective single-center cohort analysis was conducted using data from electronic medical record from 2018 to 2020. A total of 705 patients undergoing coronary artery bypass graft surgery and/or valve surgery, and/or aortic replacement surgery were included in the analysis. Intraoperative GV was assessed with a coefficient of variation (CV), which was defined as the standard deviation of five intraoperative blood glucose measurements divided by the mean. POD assessment was performed three times a day in the ICU and twice a day in the ward until discharge by trained medical staff. POD was diagnosed if any of the Confusion Assessment Method for the Intensive Care Unit was positive in the ICU, and the Confusion Assessment Method was positive in the ward. Multivariable logistic regression was used to identify associations between intraoperative GV and POD. RESULTS: POD occurred in 306 (43.4%) patients. When intraoperative glycemic CV was compared as a continuous variable, the delirium group had higher intraoperative glycemic CV than the non-delirium group (22.59 [17.09, 29.68] vs 18.19 [13.00, 23.35], p < 0.001), and when intraoperative glycemic CV was classified as quartiles, the incidence of POD increased as intraoperative glycemic CV quartiles increased (first quartile 29.89%; second quartile 36.67%; third quartile 44.63%; and fourth quartile 62.64%, p < 0.001). In the multivariable logistic regression model, patients in the third quartile of intraoperative glycemic CV were 1.833 times (OR 1.833, 95% CI: 1.132-2.967, p = 0.014), and patients in the fourth quartile of intraoperative glycemic CV were 3.645 times (OR 3.645, 95% CI: 2.235-5.944, p < 0.001) more likely to develop POD than those in the first quartile of intraoperative glycemic CV. CONCLUSION: Intraoperative blood glucose fluctuation, manifested by intraoperative GV, is associated with POD after cardiac surgery. Patients with a higher intraoperative GV have an increased risk of POD.

Cephalad misplacement of a pulmonary artery catheter in a patient with a preexisting Hickman catheter.

BACKGROUND: Pulmonary artery catheter insertion is a routine practice in high-risk patients undergoing cardiac surgery. However, pulmonary artery catheter insertion is associated with numerous complications that can be devastating to the patient. Incorrect placement is an overlooked complication with few case reports to date. CASE PRESENTATION: An 18-year-old male patient underwent elective mitral valve replacement due to severe mitral valve regurgitation. The patient had a history of synovial sarcoma, and Hickman catheter had been inserted in the right internal jugular vein for systemic chemotherapy. We made multiple attempts to position the pulmonary artery catheter in the correct position but failed. A chest radiography revealed that the pulmonary artery catheter was bent and pointed in the cephalad direction. Removal of the pulmonary artery catheter was successful, and the patient was discharged 10 days after the surgery without complications. CONCLUSIONS: To prevent misplacement of the PAC, clinicians should be aware of multiple risk factors in difficult PAC placement, and be prepared to utilize adjunctive methods, such as TEE and fluoroscopy.

Anti-Obesity Effect of Pine Needle Extract on High-Fat Diet-Induced Obese Mice.

BACKGROUND: Obesity due to an excessive intake of nutrient disturbs the hypothalamus-mediated energy metabolism subsequently develops metabolic disorders. In this study, we investigated the effect of pine needle extract (PNE) on the hypothalamic proopiomelanocortin (POMC) neurons involved in the regulation of energy balance via melanocortin system and fat tissue metabolism. METHODS: We performed electrophysiological and immunohistochemical analyses to determine the effect of PNE on POMC neurons. Mice were fed a normal or high-fat diet for 12 weeks, then received PNE for the last 2 weeks to measure the following physiological indices: Body weight, food intake, fat/lean mass, glucose metabolism, and plasma leptin levels. In addition, changes of thermogenic, lipolytic, and lipogenetic markers were evaluated in brown adipose tissue (BAT) and white adipose tissue (WAT) by western blotting, respectively. RESULTS: PNE increased hypothalamic POMC neuronal activity, and the effect was abolished by blockade of melanocortin 3/4 receptors (MC3/4Rs). PNE decreased body weight, fat mass, plasma leptin levels, and improved glucose metabolism after high-fat-induced obesity. However, PNE did not change the expression of thermogenic markers of the BAT in HFD fed groups, but decreased only the lipogenetic markers of WAT. This study suggests that PNE has a potent anti-obesity effect, inhibiting lipogenesis in WAT, even though HFD-induced leptin resistance-mediated disruption of POMC neuronal activity.

Comparison of the protective effect of cytosolic and mitochondrial Peroxiredoxin 5 against glutamate-induced neuronal cell death.

Objectives: Although glutamate is an essential factor in the neuronal system, excess glutamate can produce excitotoxicity. We previously reported that Peroxiredoxin 5 (Prx5) protects neuronal cells from glutamate toxicity via its antioxidant effects. However, it is unclear whether cytosolic or mitochondrial Prx5 provides greater neuroprotection. Here, we investigated differences in the neuroprotective effects of cytosolic and mitochondrial Prx5.Methods: We analyzed patterns of cytosolic and mitochondrial H2O2 generation in glutamate toxicity using HyPer protein. And then, we confirmed the change of intracellular ROS level and apoptosis with respective methods. The mitochondrial dynamics was assessed with confocal microscope imaging and western blotting.Results: We found that the level of mitochondrial H2O2 greatly increased compared to cytosolic H2O2 and it affected cytosolic H2O2 generation after glutamate treatment. In addition, we confirmed that mitochondrial Prx5 provides more effective neuroprotection than cytosolic Prx5.Discussion: Overall, our study reveals the mechanisms of cytosolic and mitochondrial ROS in glutamate toxicity. Our findings suggest that mitochondrial ROS and Prx5 are attractive therapeutic targets and that controlling these factors be useful for the prevention of neurodegenerative diseases.

The enhancer activity of long interspersed nuclear element derived microRNA 625 induced by NF-κB.

Transposable elements (TEs) are DNA sequences that cut or introduced into the genome, and they represent a massive portion of the human genome. TEs generate a considerable number of microRNAs (miRNAs) are derived from TEs (MDTEs). Numerous miRNAs are related to cancer, and hsa-miRNA-625 is a well-known oncomiR derived from long interspersed nuclear elements (LINEs). The relative expression of hsa-miRNA-625-5p differs in humans, chimpanzees, crab-eating monkeys, and mice, and four primers were designed against the 3'UTR of GATAD2B to analyze the different quantities of canonical binding sites and the location of miRNA binding sites. Luciferase assay was performed to score for the interaction between hsa-miRNA-625 and the 3'UTR of GATAD2B, while blocking NF-κB. In summary, the different numbers of canonical binding sites and the locations of miRNA binding sites affect gene expression, and NF-κB induces the enhancer activity of hsa-miRNA-625-5p by sharing the binding sites.

Delayed Lesions on Diffusion-Weighted Imaging in Initially Lesion-Negative Stroke Patients.

BACKGROUND AND PURPOSE: Lesions on diffusion-weighted imaging (DWI) occasionally appear on follow-up magnetic resonance imaging (MRI) among initially DWI-negative but clinically suspicious stroke patients. We established the prevalence of positive conversion in DWI-negative stroke and determined the clinical factors associated with it. METHODS: This retrospective, observational, single-center study included 5,271 patients hospitalized due to stroke/transient ischemic attack (TIA) in a single university hospital during 2010 to 2017. Patients without initial DWI lesions underwent follow-up DWI imaging as a routine practice. Adjusted hazard ratios (aHRs) for recurrent stroke risk according to positive conversion were determined using Cox proportional hazard regression. Adjusted odds ratios (aORs) and 95% confidence intervals (CIs) for positive conversion among initially DWI-negative patients were estimated. RESULTS: In total, 694 (13.2%) patients (mean±standard deviation age, 62.9±13.7 years; male, 404 [58.2%]) were initially DWI-negative. Among them, 22.5% had positive-conversion on follow-up DWI. Positive conversion was associated with a higher risk of recurrent stroke (aHR, 3.12; 95% CI, 1.56 to 6.26). Early neurologic deterioration (aOR, 15.1; 95% CI, 5.71 to 47.66), atrial fibrillation (aOR, 6.17; 95% CI, 3.23 to 12.01), smoking (aOR, 3.76; 95% CI, 2.19 to 6.63), pre-stroke dependency (aOR, 1.62; 95% CI, 1.15 to 2.27), objective hemiparesis (aOR, 4.39; 95% CI, 1.90 to 10.32), longer symptom duration (aOR, 2.17; 95% CI, 1.57 to 3.08), high cholesterol (aOR, 4.70; 95% CI, 1.78 to 12.77), National Institutes of Health Stroke Scale score (aOR, 1.44; 95% CI, 1.08 to 1.91), and high systolic blood pressure (aOR, 1.01; 95% CI, 1.00 to 1.02) were associated with a higher incidence of lesions with delayed appearance. Regarding the location of lesions on follow-up DWI, 34.6% and 21.2% were in the cortex and brainstem, respectively. CONCLUSIONS: In DWI-negative stroke/TIA, positive conversion is associated with a higher risk of recurrent stroke. DWI-negative stroke with factors related to positive conversion may require follow-up MRI for a definitive diagnosis.

Sense-oriented AluYRa1 elements provide a lineage-specific transcription environment for polyadenylation.

Transposable elements cause alternative splicing (AS) in different ways, contributing to transcript diversification. Alternative polyadenylation (APA), one of the AS events, is related to the generation of mRNA isoforms in 70% of human genes. In this study, we tried to investigate AluYRa1s located at the terminal region of cynomolgus monkey genes, utilizing both computational analysis and molecular experimentation. We found that ten genes had AluYRa1 at their 3' end, and nine of these AluYRa1s were sense-oriented. Furthermore, in seven genes, AluYRa1s were expected to have a similar consensus sequence for polyadenylation cleavage. Additional computational analysis using the annotation files from the UCSC database showed that AluYRa1 was more involved in polyadenylation than in open reading frame exon splicing. To examine the extent of AluYRa1 involvement in polyadenylation, RNA-seq data from 30 normal cynomolgus monkeys were analyzed using TAPAS, a recently devised software that detects all the promising polyadenylation sites including APA sites. We observed that approximately 74% of possible polyadenylation sites in the analyzed genes were provided by sense-oriented AluYRa1. In conclusion, AluYRa1 is an Old-World monkey-specific TE, and its sense-oriented insertion at the 3'UTR region tends to provide a favorable environment for polyadenylation, diversifying gene transcripts.

Isoliquiritigenin Reduces LPS-Induced Inflammation by Preventing Mitochondrial Fission in BV-2 Microglial Cells.

Excessive microglial cell activation in the brain can lead to the production of various neurotoxic factors (e.g., pro-inflammatory cytokines, nitric oxide) which can, in turn, initiate neurodegenerative processes. Recent research has been reported that mitochondrial dynamics regulate the inflammatory response of lipopolysaccharide (LPS). Isoliquiritigenin (ISL) is a compound found in Glycyrrhizae radix with anti-inflammatory and antioxidant properties. In this study, we investigated the function of ISL on the LPS-induced pro-inflammatory response in BV-2 microglial cells. We showed that ISL reduced the LPS-induced increase in pro-inflammatory mediators (e.g., nitric oxide and pro-inflammatory cytokines) via the inhibition of ERK/p38/NF-κB activation and the generation of reactive oxygen species (ROS). Furthermore, ISL inhibited the excessive mitochondrial fission induced by LPS, regulating mitochondrial ROS generation and pro-inflammatory response by suppressing the calcium/calcineurin pathway to dephosphorylate Drp1 at the serine 637 residue. Interestingly, the ISL pretreatment reduced the number of apoptotic cells and levels of cleaved caspase3/PARP, compared to LPS-treated cells. Our findings suggested that ISL ameliorated the pro-inflammatory response of microglia by inhibiting dephosphorylation of Drp1 (Ser637)-dependent mitochondrial fission. This study provides the first evidence for the effects of ISL against LPS-induced inflammatory response related and its link to mitochondrial fission and the calcium/calcineurin pathway. Consequently, we also identified the protective effects of ISL against LPS-induced microglial apoptosis, highlighting the pharmacological role of ISL in microglial inflammation-mediated neurodegeneration.

Actin-Membrane Release Initiates Cell Protrusions.

Despite the well-established role of actin polymerization as a driving mechanism for cell protrusion, upregulated actin polymerization alone does not initiate protrusions. Using a combination of theoretical modeling and quantitative live-cell imaging experiments, we show that local depletion of actin-membrane links is needed for protrusion initiation. Specifically, we show that the actin-membrane linker ezrin is depleted prior to protrusion onset and that perturbation of ezrin's affinity for actin modulates protrusion frequency and efficiency. We also show how actin-membrane release works in concert with actin polymerization, leading to a comprehensive model for actin-driven shape changes. Actin-membrane release plays a similar role in protrusions driven by intracellular pressure. Thus, our findings suggest that protrusion initiation might be governed by a universal regulatory mechanism, whereas the mechanism of force generation determines the shape and expansion properties of the protrusion.

Longitudinal profiling of the blood transcriptome in an African green monkey aging model.

African green monkeys (AGMs, Chlorocebus aethiops) are Old World monkeys which are used as experimental models in biomedical research. Recent technological advances in next generation sequencing are useful for unraveling the genetic mechanisms underlying senescence, aging, and age-related disease. To elucidate the normal aging mechanisms in older age, the blood transcriptomes of nine healthy, aged AGMs (15‒23 years old), were analyzed over two years. We identified 910‒1399 accumulated differentially expressed genes (DEGs) in each individual, which increased with age. Aging-related DEGs were sorted across the three time points. A major proportion of the aging-related DEGs belonged to gene ontology (GO) categories involved in translation and rRNA metabolic processes. Next, we sorted common aging-related DEGs across three time points over two years. Common aging-related DEGs belonged to GO categories involved in translation, cellular component biogenesis, rRNA metabolic processes, cellular component organization, biogenesis, and RNA metabolic processes. Furthermore, we identified 29 candidate aging genes that were upregulated across the time series analysis. These candidate aging genes were linked to protein synthesis. This study describes a changing gene expression pattern in AGMs during aging using longitudinal transcriptome sequencing. The candidate aging genes identified here may be potential targets for the treatment of aging.