gebhart/patrepeval huggingface dataset:
Drag-and-drop genome insertion without DNA cleavage with CRISPR-directed integrases (2021)

Abstract Programmable and multiplexed genome integration of large, diverse DNA cargo independent of DNA repair remains an unsolved challenge of genome editing. Current gene integration approaches require double-strand breaks that evoke DNA damage responses and rely on repair pathways that are inactive in terminally differentiated cells. Furthermore, CRISPR-based approaches that bypass double stranded breaks, such as Prime editing, are limited to modification or insertion of short sequences. We present Programmable Addition via Site-specific Targeting Elements, or PASTE, which achieves efficient and versatile gene integration at diverse loci by directing insertion with a CRISPR-Cas9 nickase fused to both a reverse transcriptase and serine integrase. Without generating double stranded breaks, we demonstrate integration of sequences as large as ∼36 kb with rates between 10-50% at multiple genomic loci across three human cell lines, primary T cells, and quiescent non-dividing primary human hepatocytes. To further improve PASTE, we discover thousands of novel serine integrases and cognate attachment sites from metagenomes and engineer active orthologs for high-efficiency integration using PASTE. We apply PASTE to fluorescent tagging of proteins, integration of therapeutically relevant genes, and production and secretion of transgenes. Leveraging the orthogonality of serine integrases, we engineer PASTE for multiplexed gene integration, simultaneously integrating three different genes at three genomic loci. PASTE has editing efficiencies comparable to or better than those of homology directed repair or non-homologous end joining based integration, with activity in non-dividing cells and fewer detectable off-target events. For therapeutic applications, PASTE can be delivered as mRNA with synthetically modified guides to programmably direct insertion of DNA templates carried by AAV or adenoviral vectors. PASTE expands the capabilities of genome editing via drag-and-drop gene integration, offering a platform with wide applicability for research, cell engineering, and gene therapy. One Sentence Summary A new technology combining CRISPR-mediated genome editing and site-specific integrases enables efficient programmable gene integration at any targeted genomic locus without double-strand DNA breaks, leading to broad applications in basic science research, cell engineering, and gene therapy.
[Effect of ethanol and lipid peroxidation on testosterone biosynthesis by interstitial cells of rat testes]. (1994)

The induction of lipid peroxidation in interstitial cells of rat testes of ADP-Fe3+ complex results in the decrease of testosterone biosynthesis. Ethanol at 50 mM concentration enhances the level of malonic dialdehyde and inhibits the steroidogenesis. Effects of alcohol and ADP-Fe3+ complex both on lipid peroxidation or testosterone biosynthesis are additive. This agrees with the idea about the mediation of ethanol effects in testes by the induction of lipid peroxidation.
Effects of nontoxic lipid A and endotoxin on resistance of mice to Toxoplasma gondii. (1988)

Mice were administered nontoxic monophosphoryl lipid A (MPL) or refined standard bacterial endotoxin (RSE) prior to, simultaneously with, or after infection with Toxoplasma gondii. MPL and RSE given before or with the toxoplasma infection induced significant resistance. Administration of toxic RSE, but not nontoxic MPL, after the establishment of toxoplasma infection significantly shortened the survival time of mice.
Genetic evidence of branching in the isoprenoid pathway for the production of isopentenyl diphosphate and dimethylallyl diphosphate in Escherichia coli (2000)

An alternative mevalonate-independent pathway for isoprenoid biosynthesis has been recently discovered in eubacteria (including Escherichia coli) and plant plastids, although it is not fully elucidated yet. In this work, E. coli cells were engineered to utilize exogenously provided mevalonate and used to demonstrate by a genetic approach that branching of the endogenous pathway results in separate synthesis of the isoprenoid building units isopentenyl diphosphate (IPP) and its isomer dimethylallyl diphosphate (DMAPP). In addition, the IPP isomerase encoded by the idi gene was shown to be functional in vivo and to represent the only possibility for interconverting IPP and DMAPP in this bacterium.
Effect of standardizedAmlaextract on atherosclerosis and dyslipidemia (2006)

Emblica officinalis , commonly known as Indian gooseberry ( Amla ), is found to be effective for the reversal of dyslipidemia and intima-media thickening and plaque formation in the aorta in hypercholesterolaemic rabbits. In this study, cholesterol powder (100 mg/kg body weight) was administered orally to healthy NZ white rabbits for 4 mo to induce hypercholesterolaemia; and thereafter, amla extract was given in two doses (10 mg and 20 mg/kg/d orally) for 4 mo. Fasting lipid profile was done monthly and also at the end of treatment. After sacrificing the animals, tissue cholesterol (liver, heart and kidney) and 3-hydroxy-3-methylglutaryl-Coenzyme A reductase activity of liver were estimated and part of aorta and myocardium were processed for histological studies. Feeding of amla extract (10 mg and 20 mg/kg) for 4 mo reversed these changes and the lumen of the aorta became normal as in the normal control group. Reversal of dyslipidemia and atheromatous plaques achieved by amla extract seems to be brought about by a number of factors, such as its ability to prevent low-density lipoprotein oxidation, its antioxidant action, besides decreasing synthesis of cholesterol by inhibiting 3-hydroxy-3-methylglutaryl-Coenzyme A reductase activity and elevating high-density lipoprotein level to enhance reverse cholesterol transport.
The case for frequency domain PD testing in the context of distribution cable (2003)

Frequency domain PD testing offers a rich set of PD characterization tools, the most basic of which are described in the article. PD detection sensitivity under field conditions is one to two orders of magnitude greater than for time domain testing as a result of (i) no need to trigger on the first PD pulse above the broadband noise, and (ii) the filtering effect of the cable between the PD detection site and the terminations. As a result of this greatly increased sensitivity and the rich set of characterization tools, frequency domain PD testing has been developed into a highly sensitive and reliable tool for characterizing the condition of distribution cable during normal operation, the sensitivity and accuracy of which have been confirmed through independent testing.
Reduced frequency of FOXP3+ CD4+CD25+ regulatory T cells in patients with chronic graft-versus-host disease (2005)

Chronic graft-versus-host disease (cGVHD) is a major complication of allogeneic hematopoietic stem cell transplantation but the immune mechanisms leading to the diverse clinical manifestations of cGVHD remain unknown. In this study, we examined regulatory T cells (Tregs) in 57 transplant recipients (30 with cGVHD and 27 without active cGVHD) and 26 healthy donors. Phenotypic studies demonstrated decreased frequency of CD4+CD25+ T cells in patients with cGVHD compared with patients without cGVHD (P < .001) and healthy individuals (P < .001). Gene expression of Treg transcription factor FOXP3 was reduced in cGVHD patients compared with patients without cGVHD (P = .009) or healthy donors (P = .01). T-cell receptor excision circle (TREC) assays for the evaluation of thymus activity revealed fewer TRECs in both transplant groups compared with healthy donors (P < .001 and P = .02, respectively) although no difference was observed between patients with or without cGVHD (P = .13). When tested in functional assays, Tregs from both patient cohorts and healthy individuals mediated equivalent levels of suppression. Collectively, these studies indicate that patients with active cGVHD have reduced frequencies of Tregs but the function of these cells remains normal. These findings support the development of new strategies to increase the number of Tregs following allogeneic hematopoietic stem cell transplantation to prevent or correct cGVHD.
Saccharomyces cerevisiae Flap Endonuclease 1 Uses Flap Equilibration To Maintain Triplet Repeat Stability (2004)

Flap endonuclease 1 (FEN1) is a central component of Okazaki fragment maturation in eukaryotes. Genetic analysis of Saccharomyces cerevisiae FEN1 (RAD27) also reveals its important role in preventing trinucleotide repeat (TNR) expansion. In humans such expansion is associated with neurodegenerative diseases. In vitro, FEN1 can inhibit TNR expansion by employing its endonuclease activity to compete with DNA ligase I. Here we employed two yeast FEN1 nuclease mutants, rad27-G67S and rad27-G240D, to further define the mechanism by which FEN1 prevents TNR expansion. Using a yeast artificial chromosome system that can detect both TNR instability and fragility, we demonstrate that the G240D but not the G67S mutation increases both the expansion and fragility of a CTG tract in vivo. In vitro, the G240D nuclease is proficient in cleaving a fixed nonrepeat double flap; however, it exhibits severely impaired cleavage of both nonrepeat and CTG-containing equilibrating flaps. In contrast, wild-type FEN1 and the G67S mutant exhibit more efficient cleavage on an equilibrating flap than on a fixed CTG flap. The degree of TNR expansion and the amount of chromosome fragility observed in the mutant strains correlate with the severity of defective flap cleavage in vitro. We present a model to explain how flap equilibration and the unique tracking mechanism of FEN1 can collaborate to remove TNR flaps and prevent repeat expansion.
Targeting plasmid‐encoded proteins to the antigen presentation pathways (2004)

The antigen presentation pathways constitute a fulcrum on which adaptive immunity is balanced, and their manipulation should allow us to induce designer immune responses. The ease and rapidity with which DNA vaccines can be constructed and altered make them ideal candidates with which to test the various targeting strategies that have been conceived to date. These approaches and the mechanisms that may (or may not) underlie their success are reviewed in this article.
Outbreak of Brucellosis at a South‐Australian Abattoir: 2. Epidemiological Investigations (1981)

The outbreak of human brucellosis among employees of a large South Australian abattoir described previously coincided with an increase in the number of cattle showing a positive serological reaction for Brucella abortus being slaughtered. Comparisons showed that two other abattoirs in the area were slaughtering larger numbers of such cattle, but no cases of human brucellosis were diagnosed there. This suggested an additional risk at the abattoir concerned. All infected men had been employed in a particular part of the works. There was a possibility of movement of aerosols, produced on opening the uteri of pregnant cattle, to other parts of the works, putting a larger number of workers at risk of infection. Modifications to the plant greatly reduced the spread of aerosols. No cases of human brucellosis were recorded at this abattoir during the summer of 1980-81.
QTL analysis of quantitative resistance toPhytophthora infestans(late blight) in tomato and comparisons with potato (2004)

Quantitative trait loci (QTLs) for resistance to Phytophthora infestans (late blight) were mapped in tomato. Reciprocal backcross populations derived from cultivated Lycopersicon esculentum x wild Lycopersicon hirsutum (BC-E, backcross to L. esculentum; BC-H, backcross to L. hirsutum) were phenotyped in three types of replicated disease assays (detached-leaflet, whole-plant, and field). Linkage maps were constructed for each BC population with RFLPs. Resistance QTLs were identified on all 12 tomato chromosomes using composite interval mapping. Six QTLs in BC-E (lb1a, lb2a, lb3, lb4, lb5b, and lb11b) and two QTLs in BC-H (lb5ab and lb6ab) were most consistently detected in replicated experiments or across assay methods. Lycopersicon hirsutum alleles conferred resistance at all QTLs except lb2a. Resistance QTLs coincided with QTLs for inoculum droplet dispersal on leaves, a trait in L. hirsutum that may contribute to resistance, and dispersal was mainly associated with leaf resistance. Some P. infestans resistance QTLs detected in tomato coincided with chromosomal locations of previously mapped R genes and QTLs for resistance to P. infestans in potato, suggesting functional conservation of resistance within the Solanaceae.
Biosynthesis and mitochondrial processing of the beta subunit of propionyl coenzyme A carboxylase from rat liver. (1983)

Propionyl-CoA carboxylase (ADP-forming) (EC 6.4.1.3), an oligomer of nonidentical subunits (alpha 4 beta 4), has been localized to the mitochondrial matrix. As a first step in examining this enzyme's biogenesis, we have investigated in vitro the cell-free, rat liver RNA-directed synthesis of the beta subunit, and its post-translational transport and processing by rat liver mitochondria. The beta subunit is synthesized as a precursor approximately 7,500 daltons larger than its mature mitochondrial counterpart. The extension segment, comprising approximately 60 amino acids, is located at the NH2 terminus of the precursor. Intact mitochondria translocate the precursor across both mitochondrial membranes, and a protease localized to the mitochondrial matrix cleaves the precursor to a polypeptide identical in size and peptide composition to the mature beta subunit.
Measurement of sugar probes in serum: an alternative to urine measurement in intestinal permeability testing (1996)

The percentage dose of lactulose and mannitol excreted in urine after oral ingestion is used as a noninvasive method of assessing small intestinal permeability. The collection of incomplete or inaccurately timed urine samples can lead to errors in estimation of sugar probe molecules. We describe an HPLC method for the simultaneous determination of lactulose and mannitol in serum after oral ingestion of test sugars. We applied the test to healthy volunteers and to subjects undergoing jejunal biopsy for suspected gluten-sensitive enteropathy. The ratio of concentrations of lactulose and mannitol in serum discriminated well between subjects with a normal biopsy and those with villous atrophy, discrimination being best at 90 min postdose. The results agree well with lactulose:mannitol ratios determined in urine (r= 0.88), and the two methods can be used interchangeably. The determination of mannitol and lactulose in serum provides an acceptable alternative to urine collection and may be particularly useful in young children. It also reduces the time spent on the investigation from 5 h to 90 min.
Phase I Study of Sequence-Selective Minor Groove DNA Binding Agent SJG-136 in Patients with Advanced Solid Tumors (2009)

SJG-136 was associated with dose-limiting VLS and hepatotoxicity when administered by short injection every 21 days. DNA damage was noted, at all dose levels studied, in circulating lymphocytes. The etiology of the observed toxicities is unclear and is the subject of further preclinical research. Alternative clinical dosing strategies are being evaluated.
New field-matching technique for low-loss splices between conventional and dispersion-flattened single-mode fibres (1988)

A new technique is reported for minimizing splice loss between conventional and dispersion-flattened single-mode fibres by slightly tapering the splice. Optimal taper ratios are calculated theoretically, considering profile-diffusion processes. Considerable loss reduction is verified experimentally.
Oligonucleotides, part 5+: synthesis and fluorescence studies of DNA oligomers d(AT)5containing adenines covalently linked at C-8 with dansyl fluorophore (1990)

The synthesis of oligodeoxynucleotides d(AT)5 in which specific adenines are linked at C-8 position with dansyl fluorophores via a variable polymethylene spacer chain are reported. This was achieved by a strategy involving prelabelling at the monomeric stage followed by solid phase assembly of oligonucleotides to obtain regiospecifically labeled oligonucleotides. Several mono and polydansyl d(AT)5 derivatives in which the fluorophore is linked via ethylene, tetramethylene and hexamethylene spacer arms were synthesised for a systematic study of their fluorescence characteristics. It was observed that (i) enhancements in fluorescence intensity and emission quantum yields are seen due to multiple labelling, (ii) the magnitude of enhancements are related to labelling configuration and (iii) quenching efficiency is minimal with shorter and rigid spacer arms. The results may aid rational design of multiple fluorescent DNA probes for nonradioactive detection of nucleic acids.
Bladder cancers respond to intravesical instillation of (HAMLET human α‐lactalbumin made lethal to tumor cells) (2007)

We studied if bladder cancers respond to HAMLET (human alpha-lactalbumin made lethal to tumor cells) to establish if intravesical HAMLET application might be used to selectively remove cancer cells in vivo. Patients with nonmuscle invasive transitional cell carcinomas were included. Nine patients received 5 daily intravesical instillations of HAMLET (25 mg/ml) during the week before scheduled surgery. HAMLET stimulated a rapid increase in the shedding of tumor cells into the urine, daily, during the 5 days of instillation. The effect was specific for HAMLET, as intravesical instillation of NaCl, PBS or native alpha-lactalbumin did not increase cell shedding. Most of the shed cells were dead and an apoptotic response was detected in 6 of 9 patients, using the TUNEL assay. At surgery, morphological changes in the exophytic tumors were documented by endoscopic photography and a reduction in tumor size or change in tumor character was detected in 8 of 9 patients. TUNEL staining was positive in biopsies from the remaining tumor in 4 patients but adjacent healthy tissue showed no evidence of apoptosis and no toxic response. The results suggest that HAMLET exerts a direct and selective effect on bladder cancer tissue in vivo and that local HAMLET administration might be of value in the future treatment of bladder cancers.
Functional integrity of the CD28 co-stimulatory pathway in T lymphocytes from elderly subjects (1999)

these experiments demonstrate the integrity of the CD28 pathway in elderly people, and suggest that ageing does not affect different T cell activation pathways equally.
[Clomiphene citrate therapy in idiopathic male infertility]. (1988)

Clomiphene citrate therapy was performed on 30 patients with idiopathic male infertility, who were intractable to non-hormonal therapy. Clomiphene citrate was administered orally at a dose of 25 mg per day for 25 days with 5 days of rest following Paulson' protocol, and we favorable results were obtained: Both sperm count and sperm motility improved in 11 cases (36.7%). Odera's score count decreased in 14 cases (46.7%), which were judged as effective. The wives of 4 patients treated by this therapy became pregnant. After this therapy, the serum levels of luteinizing hormone, follicle stimulating hormone, testosterone and prolactin were 1.90, 1.81, 1.89 and 1.22 times higher, respectively. These findings indicate that clomiphene citrate therapy is an effective for idiopathic male infertility, but we need to consider that the sudden and marked elevation of serum testosterone level by this therapy might affect male accessory sex organs such as the prostatic gland which shows androgen dependency later in life.
Therapeutic Electrical Stimulation of the Hypoglossal Nerve in Obstructive Sleep Apnea (2001)

The findings demonstrate the feasibility and therapeutic potential for hypoglossal nerve stimulation in obstructive sleep apnea.
Vulnerability to chosen-cyphertext attacks of optical encryption schemes based on double random phase keys (2005)

We show how optical encryption methods based on double random phase keys are vulnerable to an organized attack of the chosen-ciphertext type. The decryption key can be easily obtained by an opponent who has repeated access to either the encryption or decryption machines. However, we have also devised a solution that prevents the attack. Our results cast doubts on the present security of these techniques.
A Role for TGF-β in the Generation and Expansion of CD4+CD25+ Regulatory T Cells from Human Peripheral Blood (2001)

An elusive goal in transplanting organs across histocompatibility barriers has been the induction of specific tolerance to avoid graft rejection. A considerable body of evidence exists that the thymus produces regulatory T cells that suppress the response of other T cells to antigenic stimulation. We report that TGF-beta can induce certain CD4+ T cells in the naive (CD45RA+RO-) fraction in human peripheral blood to develop powerful, contact-dependent suppressive activity that is not antagonized by anti-TGF-beta or anti-IL-10 mAbs. The costimulatory effects of TGF-beta on naive CD4+ T cells up-regulated CD25 and CTLA-4 expression, increased their transition to the activated phenotype, but decreased activation-induced apoptosis. Suppressive activity was concentrated in the CD25+ fraction. These CD4+CD25+ regulatory cells prevented CD8+ T cells from proliferating in response to alloantigens and from becoming cytotoxic effector cells. Moreover, these regulatory cells exerted their suppressive activities in remarkably low numbers and maintained these effects even after they are expanded. Once activated, their suppressive properties were Ag nonspecific. Although <1% of naive CD4+ T cells expressed CD25, depletion of this subset before priming with TGF-beta markedly decreased the generation of suppressive activity. This finding suggests that CD4+CD25+ regulatory T cells induced ex vivo are the progeny of thymus-derived regulatory T cells bearing a similar phenotype. The adoptive transfer of these regulatory T cells generated and expanded ex vivo has the potential to prevent rejection of allogeneic organ grafts.
Dichotomy between naïve and memory CD4+T cell responses to Fas engagement (1999)

Engagement of Fas (APO-1, CD95), a member of the tumor necrosis factor receptor superfamily, can induce apoptotic cell death. However, Fas engagement also can costimulate lymphocyte proliferation. The physiologic regulation of these two outcomes is poorly understood. Here, we have used two systems, the first in vitro and the second in vivo, to demonstrate that naïve and memory CD4(+) T cells display dichotomous responses to Fas ligation. Naïve CD4(+) T cells (CD44(lo), CD45RB+, CD62L+) die as a consequence of Fas ligation in the presence of anti-CD3 antibody, whereas memory T cells (CD44(hi), CD45RB-, CD62L-), freshly isolated from the same starting population and subjected to the same stimulation conditions, are costimulated to proliferate by Fas ligation. In vitro, we demonstrate that CD28-mediated signals or T helper 1 and T helper 2 differentiation cytokines alter the response of naïve T cells, but not of memory T cells, to Fas ligation. In vivo experiments in hen egg lysozyme (HEL) T cell receptor transgenic mice show that CD4(+) T cells from HEL-naïve mice are killed by Fas ligation, but CD4(+) T cells from long-term HEL-exposed mice are costimulated by Fas ligation. Thus, the physiological outcome of Fas ligation in CD4(+) T cells is determined primarily by the antigenic history of the T cell.
Atrial Septal Defects in the Adult (2006)

P atients with isolated atrial septal defects (ASDs) have benefited from important recent advances in the diagnosis, evaluation, and management of their conditions. This review will focus on adolescent and adult patients with sizeable ASDs who do not have other major associated cardiac defects.